Facile and sensitive spectrophotometric determination of propoxur in formulations and environmental samples

Article abstract of DOI:10.1002/hlca.200590072

A facile, rapid, and sensitive spectrophotometric method for the determination of propoxur in insecticidal formulations, fortified water, vegetables, agricultural wastewater, and agricultural soil samples has been elaborated. The proposed method is based on the hydrolysis of propoxur under basic conditions, followed by instantaneous azo coupling of the resulting 2-isopropoxyphenol with the anilines 2a - c. This yielded the orange-red chromophore 3a (λ_max = at 470 nm). the pale-red coupling product 3b (490 nm), or the red derivative 3c (478 nm), which are stable for 46 h, 38 h, and 24 h, respectively, and could be readily analyzed spectrophotometrically.

Full text of DOI:10.1002/hlca.200590072

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Helvetica Chimica Acta ± Vol. 88 (2005)
Facile and Sensitive Spectrophotometric Determination of Propoxur in
Formulations and Environmental Samples
a
a
a
b
a
by Kailasa Suresh Kumar ), Kanchi Suvardhan ), Dasari Rekha ), B. Jayaraj ), and Pattium Chiranjeevi* )
a) Environmental Monitoring Laboratory, Department of Chemistry, S.V. University, Tirupati 517502, A.P.,
India (phone: ‡ 91-8772250556; fax: ‡ 91-8772261274; e-mail: chiranjeevi_sai@yahoo.co.in)
b
)
Department of Mathematics, S.V. University, Tirupati 517502, A.P., India
A facile, rapid, and sensitive spectrophotometric method for the determination of propoxur in insecticidal
formulations, fortified water, vegetables, agricultural wastewater, and agricultural soil samples has been
elaborated. The proposed method is based on the hydrolysis of propoxur under basic conditions, followed by
instantaneous azo coupling of the resulting 2-isopropoxyphenol with the anilines 2a ± c. This yielded the orange-
red chromophore 3a (lmax ˆ at 470 nm), the pale-red coupling product 3b (490 nm), or the red derivative 3c
(478 nm), which are stable for 46 h, 38 h, and 24 h, respectively, and could be readily analyzed spectrophoto-
metrically.
Introduction. ± Propoxur (ˆ2-isopropoxyphenyl methyl carbamate) is an insecti-
cide widely used in combating insects in agriculture and households, and public-health
pests. It provides control of insects that are resistant to chlorinated hydrocarbons and
organo phosphates. In view of its wide application, there is a need to develop new
sensitive and reliable methods for the assessment of this insecticide in H O and
2
environmental samples. Because of its significance, several methods have been
developed for the determination of propoxur: colorimetry [1][2], gas chromatography
(GC) [3 ± 6], and high-performance liquid chromatography (HPLC) [7]. However,
spectrophotometric techniques are most widely used for the determination of
insecticides because these methods are usually inexpensive and straight-forward.
Spectrophotometric techniques for the determination of propoxur are typically
based on alkaline hydrolysis, followed by coupling of the resulting phenol derivative
with different diazotised reagents such as ꢀ3-nitroaniline-4-sulphonic acidꢁ (ˆ4-amino-
2-nitrobenzenesulfonic acid) [8], ꢀ3-methyl-2-benzothiazolinone hydrazone hydro-
chlorideꢁ [9], ꢀ3,5-dibromo-p-benzoquinon chlorimineꢁ [10], ꢀp-dimethyl-phenylenedi-
amine dihydrochlorideꢁ [11], 2-aminobenzophenone [12], ꢀ4-amino-antipyrineꢁ [13],
and 4-chloroaniline [14]. Venkateswarlu and Seshaia reported an extraction-based,
sensitive spectrophotometric method for the determination of propoxur in the presence
of oxidising agent [15]. Some of the above reagents suffer from selectivity, some are not
very sensitive, and some require suitable solvents for the extraction of the
chromophore, are time consuming, or costly.
In this paper, we describe an alternative spectrophotometric method for the facile,
selective, accurate, and inexpensive determination of propoxur formulations, of
propoxur in fortified H O, vegetables, agricultural wastewater, and agricultural soil
2
samples.
ꢂ 2005Verlag Helvetica Chimica Acta AG, Zürich

Helvetica Chimica Acta ± Vol. 88 (2005)
1023
Results and Discussion. ± 1. Method. Our analytical method is based on alkaline
hydrolysis of propoxur, which gives rise to 2-isopropoxyphenol (1; Scheme). The latter
was coupled with diazotised 2,6-dibromo-4-methylbenzenamine (2a), 2,6-dibromo-4-
nitrobenzenamine (2b), or 2,4,6-tribromobenzenamine (2c) ± readily available
commercially or by bromination of the corresponding anilines ± and the resulting
azo products 3a ± c were spectrophotometrically determined in solution.
The effect of NaOH concentration on hydrolysis was studied by varying the amount
of base added (1 ± 7 ml of a 2% NaOH solution). We found that 4 ± 6 ml 2% NaOH
solution were necessary for complete hydrolysis; thus, a volume of 5ml was selected
for all experiments. Absorbance values were obtained within a pH range of 11 ± 12.
Buffer was not required to stabilise the colour. Other alkaline solutions were also
tested, but NaOH gave the best results.
Scheme
All diazotizations were carried out at 0 ± 58 in the presence of 0.5± 1.5 m HCl. In this
acidity range, constant absorbances were observed. Above this range, a decrease in the
absorbance of 3 was observed. Thus, we performed all reactions in 1m aqueous HCl,
which was optimal from both synthetic and spectral points of view. We also studied the
effect of reagent concentration (1 ± 6 ml of a 0.2% reagent solution) in 1m HCl. We
found that 4 ml of 0.2% reagent solution was sufficient for complete colour
development. Higher concentrations did not enhance the absorbance any further,
and lower concentrations gave poor results.
Possible effects of non-target species (at mg/ml levels) on the determination of
propoxur were studied. Interferences of carbaryl, carbofuran, and other non-target
pesticides, if present, could be removed by washing the CHCl extracts containing 3
3
with 0.5± 1.0% NaOH solution and with H O.
2
2
. Analytical Data. Linear calibration graphs for the spectrophotometric determi-
nation of 3 were obtained for concentrations in the range of 0.6 to 10 mg/ml. The limit
of quantification (LOQ) was determined by taking the ratio of standard deviation (s)

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024
Helvetica Chimica Acta ± Vol. 88 (2005)
of the blank with respect to H O and the slope (s) of the calibration curve multiplied by
2
a factor of ten. This means that the LOQ was ca. 4.9-times the limit of detection
(
LOD). Naturally, LOQ slightly crosses the limit of the range given by Lambert ±
Beerꢁs law; however, the LOD was still well below the corresponding lower-limit. The
upper range was determined by a plot of absorbance (at lmax) against concentration.
The calibration graph had correlation coefficients from r ˆ 0.999 to 1.009. The molar
À1
À1
absorptivity was 21800 ± 23400 l mol cm . The optical characteristics of the coloured
derivatives 3 are given in Table 1.
Table 1. Optical Characteristics, Precision, and Accuracy of the Spectrophotometric Determination of Propoxur
by Means of Chromophores 3
Characteristics
3a
3b
3c
Concentration range [mg/ml]
0.4 ± 10.0
470
0.8 ± 10.0
490
0.6 ± 10.0
478
l
max [nm]
Stability [h]
Molar absorptivity [l mol cm
Sandellꢁs sensitivity [mg cm
Limit of detection [mg/ml]
46
2.28 ´ 10
24
2.34 ´ 10
0.012
38
À1
À1
4
4
2.18 ´ 104
0.011
]
À2
]
0.013
0.2950.672
1.472
0.041
0.087
1.74
0.426
Limit of quantification [mg/ml]
3.361
0.059
0.008
1.76
2.131
0.053
0.093
1.75
a
Regression ) slope b
Regression intercept a
Rel. standard deviation [%] )
b
Correlation coefficient (r)
Rel. error [%]
0.999
1.32
1.009
1.051.28
0.999
a) Regression curve: y ˆ ax ‡ b, where x is the concentration of 3 (in mg/ml) and y is the absorbance of 3.
b
)
Determined for n ˆ 10.
3
. Propoxur Determinations. We first tested out method by determining the content
ꢃ
of propoxur in commercial insecticidal formulations: Baygon spray (1% content) and
Baygon granules (Bait; 2%). When analysing the 1% spray by means of colorimetric
ꢃ
analyses of the products 3a, 3b, and 3c, respectively, contents of 0.982 (Æ0.018),
0
.973% (Æ0.026), and 0.980 (Æ0.019) of propoxur were obtained; similarly, the 2%-
granule baits were found to contain 1.94 (Æ0.036), 1.93 (Æ0.08), and 1.90% (Æ0.09) of
insecticide, respectively.
Next, we determined the propoxur content of technical-grade samples. As shown in
Table 2, our method gave excellent results both with respect to precision (see ꢀtakenꢁ vs.
ꢀ
foundꢁ) as well as propoxur recovery (typically 99%).
Similar high levels of recovery were obtained for propoxur in both fortified water
and vegetable samples, as summarized in Table 3. Here, chromophore 3a turned out to
provide the best results, with average recoveries of 95± 99% from tap-water, distilled-
water, tomato, and potato samples. In case of 3b and 3c, somewhat more-fluctuating
values in the range of 90 ± 99.5and 93 ± 98% were found. Generally, the recoveries
decreased in the order: tap water> distilled water> potato > tomato.
4
. Statistical Analysis. The performance of the new method was compared
statistically in terms of Studentꢁs test (t-values) and the variance ratio of the ꢀF-testꢁ.
These tests showed no significant difference in accuracy and precision between the
proposed methods and reference methods [6][15]. The low values of the relative

Helvetica Chimica Acta ± Vol. 88 (2005)
1025
Table 2. Recovery of Propoxur in Technical-Grade Samples Determined Spectrophotometrically via Chromo-
phores 3
3a
3b
3c
Taken
mg/ml]
Found
[mg/ml
Recovery
[%]
Found
[mg/ml
Recovery
[%]
Found
[mg/ml]
Recovery
[%]
À1
À1
[
]
]
3
3
3
3
3
4
.50
.60
.70
.80
.90
.00
3.49
3.55
3.69
3.76
3.89
3.99
99.85
98.77
99.78
99.07
99.89
99.80
3.48
3.54
3.67
3.78
3.88
3.97
99.68
98.50
99.21
99.52
99.64
99.30
3.48
3.50
3.68
3.75
3.82
3.98
99.45
97.22
99.65
98.68
97.94
99.72
9.52 (0.48)a)
99.30 (1.02) )
a
98.77 (0.44) )
a
9
a) Average and, in parentheses, standard deviation.
Table 3. Average Recovery (in %) of Propoxur from Fortified-Water and Vegetable Samples as Determined
Spectrophotometrically from Compounds 3
Fortification [mg/ml] Tap water
Distilled water
SD
Tomato
Potato
Recovery SDa) Recovery
Recovery SD
Recovery SD
3
3
3
a
b
c
0.4 ± 2.0
0.6 ± 2.2
0.8 ± 2.4
98.48
97.52
96.23
0.59
1.69
1.42
97.44
97.03
96.15
0.31
1.59
3.06
96.79
95.47
95.50
1.24
3.52 96.85
1.77 96.50
97.64
0.38
1.32
2.94
a) Standard deviation.
standard deviation further reflect the high precision of our method. Theoretical t-values
at the 95% confidence level were found to be 2.75. A comparison of the results of our
experiments with similar, known procedures is given in Table 4. As can be seen, our
method gave satisfactory results for the determination of propoxur in water, vegetables,
and agricultural soil.
Conclusions. ± Our analytical method for the determination of the insecticide
propoxur in a variety of samples (water, vegetables, soil) was found to be facile, rapid,
inexpensive, and accurate. The coupling reagents 2a ± c are commercially available or
can be readily prepared in one step. The proposed methods do not require any special
solvent for the extraction of the chromophores 3, and the latter are quite stable relative
to other species reported in the literature [10 ± 15].
The authors are grateful to Bayer India Ltd. (Bombay, India) for the supply of technical-grade propoxur.
We also thank the head of the Dept. of Biotechnology of the S.V. University (Tirupati, India) for providing
instrumental facility to carry out this work.
Experimental Part
2
General. All chemicals were of anal. grade, and doubly-distilled H O was used throughout the experiments.
Propoxur stock soln. (1000 mg/ml) was prepared from 0.10 g of pure insecticide dissolved in 100 ml of carbonyl
free MeOH. The stock soln. was progressively diluted with the solvent to obtain standard solns. of desired
concentration. Compounds 2a or 2c (0.2 g each), dissolved in MeOH (5ml), and compound 2b in acetone

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Helvetica Chimica Acta ± Vol. 88 (2005)

Helvetica Chimica Acta ± Vol. 88 (2005)
1027
in
(
1
5ml), were diluted with 100 ml of H
HCl, were used, respectively. Absorbance measurements were performed on a Hitachi U-2001
spectrophotometer in a 1-cm quartz cell at r.t.
2
O. For hydrolysis and diazotisations, 2% aq. NaOH, and 0.3% NaNO
2
m
Coupling Reagents. All reagents 2 are commercially available (Aldrich or Fluka). They can also be readily
prepared by bromination of the corresponding anilines. General procedure for the synthesis of 2,6-dibromo-4-
methylbenzenamine (2a): To a soln. of 4-methylbenzeneamine (2 g) in 1:1 HCl/H
2
O (25ml), a soln. of Br
2
(2 ml) in AcOH (5ml) was added dropwise while stirring, the temp. being kept below 38 8. After 3 h, the
2
precipitate was filtered, washed with H O, and dried at 408 to afford 2a (80%). M.p. 75± 77 8. Compounds 2b and
2
c were prepared analogously, and recrystallized from aq. MeOH.
Sample Preparation. a) The fortified H O samples containing 0.4 ± 2.4 mg/ml of propoxur (Table 3) were
extracted with CHCl , the combined org. extracts were washed with 0.1m K CO soln. to break any emulsion
formed during the extraction, and dried (Na SO ). Finally, the solvent was evaporated, and the dry residue was
2
3
2
3
2
4
dissolved in MeOH from which the amount of insecticide was determined spectrophotometrically. b) In the case
of vegetables, 50 g of fresh vegetable or fruit was uniformly mixed with MeOH (10 ml) containing propoxur
(
10 mg), and left standing for 24 h. This material was extracted according to a procedure described in [9]. The
extracts were evaporated on a steam bath, and the residue was dissolved in MeOH and analyzed
spectrophotometrically. c) Agricultural wastewater samples (10 ml) were extracted with CHCl
3
(2 Â 10 ml).
The org. extract was evaporated to dryness at 51 Æ 68 under reduced pressure, the residue was dissolved in 50%
aq. MeOH (25ml), and suitable aliquots were analysed spectrophotometrically. d) Different soil samples (10 g)
were treated and analysed as outlined earlier [13].
General Procedure for the Determination of Propoxur. Propoxur test soln. (20 ml) was hydrolyzed for 5min
with 2% NaOH soln. (5ml) at r.t. The diazonium salts of compounds 2 were prepared by mixing a 0.3% soln. of
NaNO
2
(2 ml) with a 0.2% soln of 2 (4 ml) and 1m aq. HCl (2 ml). Coupling between diazotised 2 and the
hydrolysate at 08 yielded the chromophores 3.
REFERENCES
1] A. Ambrus, E. Hargitan, G. Karoly, A. Fulop, J. Lantos, J. Assoc. Off. Anal. Chem. 1981, 64, 743.
2] K. M. Appaiah, U. C. Nag, J. Puranaiak, V. Nagaraja, O. P. Kapur, Indian Food Packer 1984, 38, 28.
3] H. A. Moye, J. Agric. Food Chem. 1975, 23, 415.
4] I. Fogy, E. R. Schmid, J. F. K. Huber, Z. Lebensm. Unters. Forsch. 1980, 170, 194.
5] K. M. Appaiah, O. P. Kapur, K. V. Nagaraja, J. Assoc. Off. Anal. Chem. 1983, 66, 105.
6] T. H. Kim, W. C. Park, J. H. Park, T. Chung, Y. T. Lee, Bull. Korean Chem. Soc. 2003, 24, 11.
7] M. Ramaswamy, Pestic. Sci. 1974, 5, 12, 383.
8] G. Mukherjee, A. J. Mukherjee, B. R. Roy, J. Food Sci. Tech. 1975, 12, 96.
9] C. S. P. Sastry, D. Vijaya, Talanta 1987, 34, 372.
[
[
[
[
[
[
[
[
[
[
[
[
[
[
[
10] W. F. Vangils, Analyst 1979, 104, 1185.
11] C. S. P. Sastry, D. Vijaya, D. S. Mangala, Analyst 1987, 112, 75 .
12] D. Venkaiah, P. R. Naidu, Talanta 1990, 37, 629.
13] P. Venkateswarlu, V. Chandra Kumar, K. Seshaiah, Ind. J. Chem., Sect. B 1995, 34, 67.
14] V. Harikrishna, N. V. S. Naidur, Asian J. Chem. 2001, 13, 687.
15] P. Venkateswarlu, K. Seshaiah, Talanta 1994, 42, 73.
Received December 21, 2004