Distribution and elimination of 14C-ethion insecticide in chamomile flowers and oil

Article abstract of DOI:10.1080/10426507.2011.588506

The residual fate of ¹⁴C-ethion labeled at the ethyl group in chamomile flowers and oil was studied. ¹⁴C-ethion and some of its degradation products have been prepared for the present investigation. ¹⁴C-residues in its flowers and oil were determined at different time intervals. Chromatographic analysis of chamomile oil extracts revealed the presence of the parent compound together with four metabolites, which were identified as ethion monooxon, ethion dioxon, O,O-diethyl phosphorothioate, and O-ethyl phosphorothioate. Effect of drying on ethion residues in chamomile flowers was investigated. Ethion residues were decreased by sun drying of chamomile flowers. The percentage of removal was 18%-47%. Effect of using low cost adsorbents for removal of ethion residues from chamomile oil was investigated. The removal percentage due to adsorption was 100% for CaO and saw dust adsorbers and for watermelon peels, bagass, and rice bran adsorbers were 36%, 41%, and 90%, respectively. Our results proved that the addition of CaO saw dust and rice bran reduced the pesticide residues, so we recommend the addition of these adsorbents during the extraction process to reduce the risk of pesticide residues on chamomile oil. Copyright Taylor & Francis Group, LLC.

Full text of DOI:10.1080/10426507.2011.588506

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Distribution and Elimination of 14C-
Ethion Insecticide in Chamomile Flowers
and Oil
H. Abdel-Gawad a , R. M. Abdelhameed a , A. M. Elmesalamy b & B.
Hegazi a
a Applied Organic Chemistry Department , NRC , Dokki, Cairo, Egypt
b Chemistry Department, Faculty of Science , Zagazig University ,
Zagazig, Egypt
Published online: 14 Oct 2011.
To cite this article: H. Abdel-Gawad , R. M. Abdelhameed , A. M. Elmesalamy & B. Hegazi (2011)
Distribution and Elimination of 14C-Ethion Insecticide in Chamomile Flowers and Oil, Phosphorus,
Sulfur, and Silicon and the Related Elements, 186:10, 2122-2134, DOI: 10.1080/10426507.2011.588506
To link to this article: http://dx.doi.org/10.1080/10426507.2011.588506
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Phosphorus, Sulfur, and Silicon, 186:2122–2134, 2011
Copyright ꢀC Taylor & Francis Group, LLC
ISSN: 1042-6507 print / 1563-5325 online
DOI: 10.1080/10426507.2011.588506
DISTRIBUTION AND ELIMINATION OF 14C-ETHION
INSECTICIDE IN CHAMOMILE FLOWERS AND OIL
H. Abdel-Gawad,1 R. M. Abdelhameed,1 A. M. Elmesalamy,2
and B. Hegazi1
1Applied Organic Chemistry Department, NRC, Dokki, Cairo, Egypt
2Chemistry Department, Faculty of Science, Zagazig University, Zagazig, Egypt
GRAPHICAL ABSTRACT
OC2H5
OC2H5
S
S
P
C2H5
C2H5
O
S
S
C
H2
P
(II)
O
oxidation
hydrolysis
OC2H5
OC2H5
O
P
S
OC2H5
OC2H5
C2H5
C2H5
O
S
C
S
HO
P
(V)
(III)
H2
P
S
O
hydrolysis
hydrolysis
S
oxidation
OH
OC2H5
OC2H5
C2H5
C2H5
O
O
S
S
C
OH
HO
P
H2
P
OC2H5
P
(VIII)
C2H5
C2H5
O
S
C
S
O
(VII)
H2
P
Further metabolism,
conjugation
O
O
(IV)
chamomile plant
Abstract The residual fate of 14C-ethion labeled at the ethyl group in chamomile flow-
ers and oil was studied. 14C-ethion and some of its degradation products have been pre-
pared for the present investigation. 14C-residues in its flowers and oil were determined at
different time intervals. Chromatographic analysis of chamomile oil extracts revealed the
presence of the parent compound together with four metabolites, which were identified as
ethion monooxon, ethion dioxon, O,O-diethyl phosphorothioate, and O-ethyl phosphoroth-
ioate. Effect of drying on ethion residues in chamomile flowers was investigated. Ethion
residues were decreased by sun drying of chamomile flowers. The percentage of removal
was 18%–47%. Effect of using low cost adsorbents for removal of ethion residues from
chamomile oil was investigated. The removal percentage due to adsorption was 100% for
CaO and saw dust adsorbers and for watermelon peels, bagass, and rice bran adsorbers
were 36%, 41%, and 90%, respectively. Our results proved that the addition of CaO saw dust
and rice bran reduced the pesticide residues, so we recommend the addition of these adsor-
bents during the extraction process to reduce the risk of pesticide residues on chamomile oil.
Received 8 February 2011; accepted 5 May 2011.
Address correspondence to H. Abdel-Gawad, Applied Organic Chemistry Department, National Research
Centre (NRC), Dokki, Cairo 12622, Egypt. E-mail: abdelgawadhassan@hotmail.com
2122

DISTRIBUTION AND ELIMINATION OF 14C-ETHION INSECTICIDE
2123
[Supplemental materials are available for this article. Go to the publisher’s online edition of
Phosphorus, Sulfur, and Silicon and the Related Elements for the following free supplemental
resource: Figure S1: 1H NMR of Compound (IV)]
Keywords Ethion; chamomile; removal; calcium oxide; rice bran
INTRODUCTION
Chamomile (Matricaria chamomilla) flower heads and extracts are used in the phar-
maceutical industry as cosmetic repairs1 for their antispasmodic, anti-inflammatory, and
antimicrobial properties2–4 and also due to natural hair dye and fragrance.5 The organophos-
phorus pesticides most commonly used in the cultivation of essential oil plants travel through
the vascular system of the plant and are absorbed at the cellular level.6
Ethion insecticide [S,Sꢁ-methylene bis (O,Oꢁ-diethyl phosphorodithioate)] is an
organothiophosphate pesticide that was first registered for use in the United States in
1965.7 This pesticide was first developed as a nonsystemic insecticide and acaricide for use
on fruit trees, including citrus fruits, nut trees, cotton and seed, and forage crops as well as
a wide variety of fruits and vegetables.8 It is also used for control of aphids, spider mites,
scale insects, thrips, lepidopterous larvae, leafhoppers, maggots, suckers, and soil-dwelling
insects on a wide variety of food, fiber, and ornamental crops, including grapes, fruits,
vegetables, and nuts.9
Information about the effect of drying on pesticide residues in herbs or other crops
is limited in literature. Natural drying and hot air drying are still the most widely used
methods to produce dried herb flakes because of their lower cost.10
Adsorption is one of the most efficient methods for removal of pollutants.11 Activated
carbon is a very efficient adsorbent for removing varieties of pesticides due to its high surface
area and porosity.12 However, because of the high cost of activated carbon, its use in the
field is restricted on economical considerations.13 Nowadays, low cost adsorbents have
been investigated as an alternative to the high cost activated carbon, for example, fly ash,13
carbon cloth,14 porous polymeric adsorbents,15 wheat residue black carbon,16 bleaching
earth,17 lignin,18 riverbed sand,19 wood char coal,20 and waste tire rubber granules.21
To the best of our knowledge, there are no studies on the fate and removal of ethion
from chamomile oil. Therefore, this study attempted to investigate the residues of ethion
on chamomile oil. In addition, a complete picture on the reduction of this compound in
flowers and oil was also evaluated.
RESULTS AND DISCUSSION
Effect of Drying
Table 1 shows the effect of drying on ethion residues on chamomile flowers after
different time intervals of the last application. The concentration of 14C-ethion activity
decreased during experimental period and reached 2.42 ppm after 30 days; the percentage
of removal because of drying was 18.2%. Cabras et al.22,23 found that drying process could
cause an important decrease in pesticide residues in apricot and raisin processed from
grapes. Juraske et al.24 noted that the degradation of pesticides is not only due to drying but
also due to other processes such as metabolism and photodegradation.

2124
H. ABDEL-GAWAD ET AL.
Table 1 Effect of drying on the residues of 14C-ethion in chamomile oil after the last application
Time of treatment∗
Sample
(7 day) ppm
(15 day) ppm
(21 day) ppm
(30 day) ppm
Flowers before drying
Flowers after drying
% removal due to drying
4.21 0.10
2.21 0.03
47.44
4.33 0.08
3.13 0.10
26.48
3.75 0.06
2.94 0.06
21.59
2.96 0.08
2.42 0.04
18.18
∗Results are expressed as mean SD for three determinations for each sample.
Distribution of 14C-ethion Residues in Chamomile Oil
The topical application of 14C-ethion on chamomile plant led to the appearance
of 14C-activity in the oil (Table 2). 14C-ethion residue in hexane and methanol extracts
increased during the first 2 weeks and then decreased rapidly at the end of experimental
period. The nonextractable residues (bound residue) show the same trend. The maximum
binding of ethion residues was 54.1% after 30 days from the last application, and the
recovery percentage ranged from 94% to 98%. Half-life (T1/2) for degradation of ethion on
chamomile flowers was found to be 3.4 days at recommended dosage. A waiting period of 15
days is suggested for safe consumption of the chamomile flowers and oil. The data obtained
are in agreement with those reported by Singh et al., on their studies, the persistence of
ethion residues on cucumber.25
Identification and Characterization of Metabolites in Chamomile Oil
Table 3 shows the Rf values and the average concentration of 14C-ethion (parent
compound and metabolites) in chamomile oil extract after 2 and 4 weeks of the second
application, in addition to the parent compound; ethion monooxon (III), ethion dioxon
(IV), O,O-diethyl phosphorothioate (V), and O-ethyl phosphorothioate (VII) were found
as free metabolites besides O,O-diethyl-S-hydroxymethyl phosphorodithioate (VII) as a
conjugated metabolite. The concentration of 14C-ethion in chamomile oil and its metabolites
was decreased at the end of experimental periods. On the contrary, compound (VIII) which
was increased at the same period due to all degradation products are hydrolysis to the
compound (VIII) at the end of experiment period.
The metabolic fate of 14C-ethion labeled at the ethyl group was studied in chamomile
oil and flowers after double application on the leaves. At 15 days and 30 days after the
second application, the parent compound and its metabolites were found in chamomile
oil. Our results demonstrated that ethion was metabolized to ethion monooxon (III) and
ethion dioxon (IV) by oxidation and to O,O-diethyl phosphorothioate (V) and O-ethyl
phosphorothioate (VIII) by hydrolysis in the plant. Hydrolysis of ethion monooxon leads
to the formation of O,O-diethyl-S-hydroxymethyl phosphorodithioate (VII), which may be
conjugated with sugar in plants (Figure 1). While the basic features of ethion metabolism
are known, detailed information is lacking. Like other organothiophosphate insecticides
(chlorpyrifos, parathion), ethion is converted via desulfuration in the liver by cytochrome
P-450 enzymes to its active oxygen analog, ethion monooxon.26 It is not known whether
ethion monooxon can then be desulfurated to ethion dioxon. Ethion monooxon is a potent
inhibitor of cholinesterases and exerts toxicity by reacting with an inhibiting neural acetyl-
cholinesterase.27 The breakdown of ethion and ethion monooxon has not been characterized

2125

2126
H. ABDEL-GAWAD ET AL.
Table 3 Amount and Rf values of 14C-ethion and its active degradation products in chamomile oil
Rf values in various solvent
systems
Metabolites
1
2
3
(15 day) ppm
(30 day) ppm
Ethion (II)
Ethion monooxon (III)
Ethion dioxon (IV)
O,O-diethyl phosphorothioate (V)
O-ethyl phosphorothioate (VIII)
0.75
0.88
0.69
0.33
0.24
0.80
0.92
0.56
0.23
0.50
0.75
0.65
0.55
0.42
0.71
0.370
0.121
0.194
0.159
0.056
0.219
0.101
0.162
0.048
0.090
System 1: Toluene: Xylene 20: 20 (v/v).
System 2: Dioxane: Xylene: Petroleum ether 10: 20: 20 (v/v/v).
System 3: n-Hexane: Ethyl acetate 98: 2 (v/v).
but is presumed to be by esterases in the blood and liver.28,29 Samples were extracted with
ethyl acetate; the aqueous and organic phases were analyzed by high-performance liquid
chromatography (HPLC). More than 99% of the urine radioactivity was in the aqueous
phase. Another sample was acidified (presumably to hydrolyze conjugates) and also ex-
tracted with ethyl acetate. Acidification converted about 30% of the radioactivity in the
aqueous phase to an organosoluble form, which may indicate that some of the products
OC2H5
OC2H5
S
S
P
C2H5
C2H5
O
S
S
C
H2
P
(II)
O
oxidation
hydrolysis
OC2H5
OC2H5
O
P
S
OC2H5
OC2H5
C2H5
C2H5
O
S
C
S
HO
P
(V)
(III)
H2
P
S
O
hydrolysis
hydrolysis
S
oxidation
OH
OC2H5
OC2H5
C2H5
C2H5
O
O
S
S
C
OH
HO
P
H2
P
OC2H5
P
(VIII)
C2H5
C2H5
O
S
C
S
O
(VII)
H2
P
Further metabolism,
conjugation
O
O
(IV)
chamomile plant
Figure 1 Metabolic pathway of ethion in chamomile oil.

DISTRIBUTION AND ELIMINATION OF 14C-ETHION INSECTICIDE
2127
Table 4 Removal of 14C-activity from chamomile plant using foliar application of calcium chloride
14C-residues (ppm∗∗∗
)
Without CaCl2
ppm
With 1.5% CaCl2
∗
∗
Sample
%
ppm
%
Flowers before drying
Flowers after drying
% removal∗∗
2.96 0.08
2.42 0.04
0
0
1.74 0.05
1.55 0.03
41
36
18
11
Oil
0.62 0.02
0
0.36 0.01
42
∗% removal related to affect of adding CaCl2.
∗∗% removal due to drying.
∗∗∗Results are expressed as mean SD for three determinations for each sample.
of ethion metabolism are present in urine as conjugates. Four to six radiolabeled metabo-
lites were detected by HPLC; none of them migrated with standards for ethion, ethion
monooxon, or ethion dioxon. None of the metabolites were specifically identified.30–32
Removal of 14C-Activity from Chamomile Plant and Oil
Foliar Application of Calcium Chloride on Chamomile Plant. The radioac-
tivity in the flowers and oil after application of 14C-ethion and calcium chloride solution
(1.5%) was presented after 30 days (Table 4). 14C-ethion residues decreased from 2.96
to 1.74 ppm due to foliar application of 1.5% CaCl2 solution on the fresh flowers and
decreased from 2.42 to 1.55 ppm on the dried flowers. The percentage of removal due to
sun drying process was 11%. 14C-ethion residues in oil extracts after sun drying process
decreased from 0.62 to 0.36 ppm due to CaCl2 addition.
By Chemical and Low Cost Adsorbent Derived From Agricultural
and Industrial Wastes
Table 5 represents the removal of 14C-activity from chamomile oil by using chemical
and low cost adsorbents derived from agricultural wastes (rice bran, watermelon peels, saw
dust, bagass). The effect of using low cost adsorbents for removal of ethion residues from
chamomile oil was investigated. The percentage removal due to adsorption was 100% for
Table 5 Removal of 14C-activity from chamomile oil using chemical and low cost adsorbent derived from
agricultural wastes
14C-residues (ppm) (Flowers after treatment with 1.5% CaCl2)∗
With
rice
bran
With
watermelon
peels
With
saw
dust
Without
adsorber
With
CaO
With
bagass
Sample
Total 14C-activity
% removal
0.360 0.035
—
0
100
0.036 0.003
90
0.210 0.042
41
0.230 0.028
36
0
100
∗Results are expressed as mean SD for three determinations for each sample.

2128
H. ABDEL-GAWAD ET AL.
0.4% CaO and 0.5% saw dust adsorber and 36%, 41%, and 90% for 0.5% watermelon
peels, 0.5% bagass, and 0.5% rice bran adsorber, respectively. The results obtained are in
parallel with the data obtained by several authors33–37 on their studies about the effect of
low cost adsorbents on removal of pesticide residues.
CONCLUSION
The results demonstrate that the level of pesticide residues in chamomile oil is
lowered, sometimes far below the recommended levels by drying or using adsorbents. The
degradation products in chamomile oil were identified as ethion monooxon, ethion dioxon,
O-ethyl phosphorothioate, and O,O-diethyl phosphorothioate. Among various methods
used for removal of pesticide from chamomile, it is suggested that factories of essential
oils should dry chamomile flowers carefully before extractions. Also, the addition of CaO,
saw dust, and rice bran reduced the pesticide residues, so we recommend the addition of
these adsorbents during the extraction process to reduce the risk of pesticide residues on
chamomile oil.
EXPERIMENTAL
Spectral Analysis
1
The H NMR spectra were acquired using a Jeol-EX (270 MHz) spectrometer;
chemical shifts δ in ppm with tetramethylsilane (TMS) as the internal standard; coupling
constants J in Hz for samples dissolved in CDCl3. The EI-MS (70 eV) spectra were recorded
with Jeol JMS-AX500 mass spectrometer. The IR spectra were recorded in potassium
bromide rolled (KBr) on Nexus 670 FTIR spectrometer (Nicolet), at the National Research
Centre. Elemental analytical data (in accord with the calculated values) were obtained from
the Microanalytical Unit, Cairo University, Giza, Egypt.
Chemicals
14C-ethion was synthesized by the reported methods with some modification as
follows.38–40
Synthesis of 14C-Ethion (Endo) S,S ꢀ-Methylene Bis (O,O ꢀ-Diethyl
Phosphorodithioate) (II)
Synthesis of O,O-Diethyl Hydrogen Phosphorodithioate (I). Anhydrous
benzene (40 mL) was stirred with P2S5 (10 g) and temperature of the mixture was maintained
around 40 ◦C. 14C-ethanol (12 mL, Sp. Act 37 MBq) was added dropwise and stirring was
continued for 3 h. The crude oil was purified on silica gel column using hexane: acetone
(98:2) for elution to leave colorless oil of 68% yield. Thin layer chromatography (TLC) on
silica gel plate showed one spot of rate of flow (Rf) (0.68) in solvent system n-hexane: ethyl
acetate 98:2. IR (KBr) νmax/cm−1, 3400 (SH), 2851, 1580 (C H aliphatic), 1022 (POC)
and 722 (P S); 1H NMR (CDCl3, 270 MHz), δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH), 2.80
(q, J = 8.5 Hz, 2H, CH2CH3), and 6.51 (s, 1H, SH); EI-MS (m/z), 186 (M+, 35.3%),
158 (68.7%), and 130 (91.5%); Anal. calcd. (%) for C4H11O2PS2 (186.23): C, 25.80; H,
5.95; P, 16.63; S, 34.44; Found (%): C, 25.75; H, 5.91; P, 15.98; S, 34.47 (Figure 2).

DISTRIBUTION AND ELIMINATION OF 14C-ETHION INSECTICIDE
2129
S
P
OC2H5
OC2H5
4
C2H5OH
p2s5
2 HS
(I)
CH2Cl2
OC2H5
OC2H5
S
S
P
C2H5
O
S
P
C
H2
(II)
S
C2H5
H2O2
O
KMnO4
OC2H5
OC2H5
O
S
buffer
hydrolysis
OC2H5
O
S
P
C2H5
O
P
S
C
C2H5
C2H5
O
H2
S
S
C
OC2H5
P
H2
P
O
C2H5
CH2OH
O
(IV)
(III)
O
C2H5
O
S
S
P
O
C2H5
(VII)
S
S
P
S
P
Cl
OC2H5
OC2H5
NaOH
2 C2H5OH
Cl
P
Cl
HO
+
Cl
OC2H5
OC2H5
(V)
O
O
O
Cl
OC2H5
OC2H5
OC2H5
OC2H5
NaOH
2 C2H5OH
Cl
P
Cl
P
HO
P
+
Cl
(VI)
Figure 2 Preparation of ethion and its degradation products.
A solution of methylene chloride (0.05 moles, 3.5 mL) and triethylamine (0.1 mole,
13.5 mL) in dry benzene (20 mL) was added dropwise to (I). The reaction mixture was
stirred at 25 ◦C for 20 min, and then refluxed for 3 h. It was filtered to remove triethylamine
hydrochloride and evaporated under vacuum to remove the solvent. The crude oil (Sp. Act.
0.199 mci/g, 7.4 MBq/g, 68% yield) was purified on a silica gel column using hexane:
acetone (98:2) for elution to leave colorless oil. TLC on silica gel plate showed one spot
of Rf 0.75 in solvent system n-hexane: ethyl acetate 98:2. IR (KBr) νmax/cm−1, 2966, 1580
(C H aliphatic), 1022 (POC), 720 (P S) and 670 (C S); 1H NMR (CDCl3, 270 MHz),
δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH), 2.80 (q, J = 8.5 Hz, 2H, CH2 CH3), and 7.30
(s, SCH2S); EI-MS (m/z), 385 (M+, 18.1%), 365 (9.1%), 231 (20.2%), and 153 (100%);
Anal. calcd. (%) for C9H22O4P2S4 (384.48): C, 28.12; H, 5.77; P, 16.11; S, 33.36; Found
(%): C, 27.99; H, 5.90; P, 16.15; S, 33.41 (Figure 2).
Synthesis of Ethion Monooxon (III). Ethion (3.8 mL, 0.01 mole) was dissolved
in acetic acid (20 mL). A solution of sulfuric acid (2 mL) in hydrogen peroxide (5 mL)
(3.5 mole of 30%) was added dropwise maintaining the temperature between 10 ◦C and 20

2130
H. ABDEL-GAWAD ET AL.
◦C. The reaction mixture was left to stir at ambient temperature for 13 h, and chloroform
(200 mL) was added to extract with distilled water (300 mL). Subsequently, the solution was
washed with 5% aqueous sodium hydrogen carbonate until neutral; then, it was dried over
anhydrous sodium sulfate and filtered, and the solvent was then removed under vacuum.
The crude oil was purified on silica gel column using hexane: acetone (98:2) for elution
to leave colorless oil of 73% yield. TLC on silica gel plate showed one spot of Rf 0.65
in solvent system n-hexane: ethyl acetate 98:2. IR (KBr) νmax/cm−1, 2966, 1580 (C H
aliphatic), 1220 (P = O), 1022 (POC), 720 (P S), and 690 (C S); 1H NMR (CDCl3, 270
MHz), δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH), 2.80 (q, J = 8.5 Hz, 2H, CH2 CH3),
and 7.30 (s, SCH2S); EI-MS (m/z), 369 (M+, 19.1%), 349 (10.3%), 215 (100%), and 137
(69.2%); Anal. calcd. (%) for C9H22O5P2S3 (368.41): C, 29.34; H, 6.02; P, 16.81; S, 26.11;
Found (%): C, 29.45; H, 5.97; P, 16.75; S, 26.31 (Figure 2).
Synthesis of Ethion Dioxon (IV). Ethion (3.85 mL, 0.01 mole) was mixed with
a solution of potassium permanganate (50 mL, 4.2 g, 0.01 mole) and conc. sulfuric acid
(3 mL) was dissolved in water (50 mL). The solution was added dropwise to the stirred
ethion mixture while maintaining the temperature between 15 ◦C and 20 ◦C. The reaction
mixture was filtered and the filtrate was treated with sodium bisulfate to remove the
excess potassium permanganate. The mixture was then saturated with sodium sulfate and
extracted with dichloromethane (100 mL). The obtained organic extracts were then dried
over anhydrous sodium sulfate and filtered. The solvent was then removed under vacuum.
The crude oil was purified on silica gel column using hexane: acetone (98:2) for elution
to leave colorless oil of 88% yield. TLC on silica gel plate showed one spot of Rf 0.55
in n-hexane: ethyl acetate 98:2. IR (KBr) νmax/cm−1, 2966, 1580 (C H aliphatic), 1240
1
(P O), 1022 (POC), and 670 (C-S); H NMR (270 MHz, CDCl3), δ = 1.24 (t, J = 8.0
Hz, 3H, CH3CH), 2.80 (q, J = 8.5 Hz, 2H, CH2 CH3), and 7.30 (s, SCH2S); EI-MS
(m/z), 353 (M+, 8.1%), 333 (29.6%), 199 (100%), and 121 (79.3%); Anal. calcd. (%) for
C9H22O6P2S2 (352.34): C, 30.68; H, 6. 29; P, 17.58; S, 18.20; Found (%): C, 30.70; H,
1
6.33; P, 17.65; S, 18.35 (Figure 2). The H NMR of compound IV is contained in the
Supplemental Materials (Figure S1).
Synthesis of O,O-Diethyl Phosphorothioate (V). A mixture of absolute
ethanol (0.01 mole, 0.6 mL) and triethylamine (0.01 mole, 1.4 mL) in dry benzene was
added dropwise to a cooled (5 ◦C–10 ◦C) solution of thiophosphoryl chloride (0.005 mole,
0.507 mL) in dry benzene during 15 min. The reaction mixture was stirred at 25 ◦C for 18
h and filtered; most of benzene was evaporated using rotary evaporator. The residues were
dissolved in sodium hydroxide solution (100 mL, 0.1 N) after extraction with diethyl ether
(100 mL), and the aqueous phase was acidified to pH 1 with a few drops of concentrated
HCl and extracted again with ether (100 mL). The ether extract was dried over anhydrous
sodium sulfate and evaporated to leave colorless oil. The crude oil was purified on silica gel
column using hexane: acetone (98:2) for elution to leave colorless oil of 68% yield. TLC
on silica gel plate showed one spot of Rf (0.42) in n-hexane: ethyl acetate 98:2. IR (KBr)
νmax/cm−1, 3440 (OH), 2851 (C H aliphatic), 1022 (POC), and 720 (P S); 1H NMR (270
MHz, CDCl3), δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH), 7.40 (s, 1H, OH), and 2.80 (q, J =
8.5 Hz, 2H, CH2CH3); EI-MS (m/z), 170 (M+, 17.8%), 142 (100%), and 114(54.6%);
Anal. calcd. (%) for C4H11O3PS (170.17): C, 28.23; H, 6. 52; P, 18.20; S, 18.84; Found
(%): C, 28.30; H, 6.60; P, 18.35; S, 18.70 (Figure 2).
Synthesis of O,O-Diethyl Phosphoric Acid (VI). A mixture of absolute ethanol
(0.01 mole, 0.6 mL) and triethylamine (0.01 mole, 1.4 mL) in dry benzene was added
dropwise to a cooled (5 ◦C–10 ◦C) solution of phosphoryl chloride (0.005 mole, 0.7 mL) in

DISTRIBUTION AND ELIMINATION OF 14C-ETHION INSECTICIDE
2131
dry benzene during 15 min. The reaction mixture was stirred at 25 ◦C for 18 h and filtered;
most of benzene was evaporated using rotary evaporator. The residues were dissolved in
sodium hydroxide solution (100 mL, 0.1 N) after extraction with diethyl ether (100 mL),
and the aqueous phase was acidified to pH 1 with a few drops of concentrated HCl and
extracted again with ether (100 mL). The ether extract was dried over anhydrous sodium
sulfate and evaporated to leave colorless oil. The crude oil was purified on a silica gel
column using hexane: acetone (98: 2) for elution to leave colorless oil of 68% yield. TLC
on silica gel plate showed one spot of Rf (0.81) in n-hexane: ethyl acetate 98:2. IR (KBr)
νmax/cm−1, 3440 (OH), 2851 (CH aliphatic), 1220 (P O), and 1022 (POC); 1H NMR (270
MHz, CDCl3), δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH2), 2.80 (q, J = 8.5 Hz, 2H, CH2CH3),
and 7.10 (s, 1H, OH); EI-MS (m/z), 154 (M+, 29.4%), 126 (79.1%), and 98 (100%); Anal.
calcd. (%) for C4H11O4P (154.10): C, 31.18; H, 7. 19; P, 20.10; Found (%): C, 30.97; H,
7.30; P, 20.23 (Figure 2).
Synthesis of O,O-Diethyl S-Hydroxymethyl Phosphorodithioate (VII). A
mixture of ethion insecticide (0.5 mL) and acetone (5 mL) was added to 10 mL of pH (5.5)
buffer solution and placed in a covered circulating water bath at 30 ◦C for 3 h. The mixture
was extracted with benzene-hexane (50:50 v/v) (50 mL). The extracted layer was dried over
anhydrous sodium sulfate and evaporated to leave colorless oil. The crude oil was purified
on silica gel column using hexane: acetone (98:2) for elution to leave colorless oil of 68%
yield. TLC on silica gel plate showed one spot of Rf (0.85) in n-hexane: ethyl acetate 98:2.
1
IR (KBr) νmax/cm−1, 3440 (OH), 2860 (CH aliphatic), 1022 (POC), and 722 (P S); H
NMR (270 MHz, CDCl3), δ = 1.24 (t, J = 8.0 Hz, 3H, CH3CH2), 2.40 (q, J = 8.5 Hz, 2H,
CH2CH3), 7.10 (s, 1H, SCH2O), and 7.60 (s, 1H, OH); EI-MS (m/z), 216 (M+, 18.6%),
188 (80.1%), 186 (65.7%), and 160 (88.3%); Anal. calcd. (%) for C5H13O3PS2 (216.26):
C, 27.77; H, 6. 06; P, 14.32; S, 29.65; Found (%): C, 28.01; H, 6.08; P, 14.25; S, 29.73
(Figure 2).
Plant Material and Experimental Design
Chamomile plants were cultivated in an isolated and controlled area at the farm
of the National Research Center located in Dokki, Cairo, Egypt. Plants were irrigated and
fertilized as in practice; seeds were planted first in small containers, 1/8 inch deep. When the
plants were big enough to handle, they were transplanted to soil including three replicates
for each treatments. The flowers were harvested on a clear morning before the sun has
drawn the valuable scent from the blossoms. Using scissors, opened heads were carefully
picked and spread on paper in a cool, dry, airy place. After the heads became papery, they
were stored in an airtight jar. At the time of blooming, the plants were divided into two
parts; the first part was treated with insecticide and CaCl2 solution (1.5%), while the second
part was only treated with 14C-ethion (4 mg/plant each time) on leaves of plants. 14C-ethion
was topically applied manually with a micropipette on healthy leaves of plants (twice, 15
days apart). Chamomile flowers were collected at, 7, 15, 21, and 30 days after the second
spray of labeled ethion. The total radioactivity of flowers (before and after drying) was
determined by digestion followed by liquid scintillation counting (LSC).
Extraction of Chamomile Flowers
Chamomile oil was extracted from its dried flowers by using a Soxhlet apparatus
for 12 h with n-hexane. After evaporation of hexane under reduced pressure, radioactivity

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H. ABDEL-GAWAD ET AL.
Figure 3 Diagram for removal of 14C-ethion insecticide from chamomile plant and oil.
in the chamomile oil was measured. The residue remaining after extraction was further
extracted with methanol (Figure 3). Aliquots of the hexane extract (oil) and the methanol
extract were used for determination of radioactivity. The remaining cake was air dried and
digested by using 1 mL Solusol (tissue and gel solubilizer), 1 mL 30% H2O2, and 70 µL
glacial acetic at 40 ◦C–50 ◦C. The radioactivity was counted using LSC.
Adsorbent Development
Chamomile flowers were treated with low cost adsorbents, i.e., rice bran, sawdust,
baggass, and watermelon peels. To improve the sorption capacity of materials, the chemical
treatment was given by washing with deionized water to remove water-soluble compounds
from the surface of materials an◦ d to develop its porous structure. The baggass was treated
with hydrogen peroxide at 60 C for 24 h to remove the adhering matter. Rice bran, rice
husk, sawdust, and watermelon peels were then reacted with 0.1 M nitric acid for 1 h
followed by soaking in methanol for 4 h to remove inorganic an◦d organic matter from
the surface of the materials and dried in an electric oven at 110 C for 1 h, powdered,
ground, and sieved to desired particle size before use. The treated materials were then
stored in desiccators to be used as sorbents. The radioactivity in cakes after oil extraction
was determined by digestion followed by LSC.
Isolation and Characterization of 14C-Residues
An analysis of the radioactive compounds was achieved by TLC. Samples of crude
oil after 2 and 4 weeks of the second application were partitioned between acetonitrile and
hexane three times to remove the oil. The radioactive residues were distributed between the
two layers. Analysis of oil extracts was achieved by TLC on silica gel plates (Merck-silica
gel 60 F254). The solvent systems used were:

DISTRIBUTION AND ELIMINATION OF 14C-ETHION INSECTICIDE
2133
• System 1: Toluene: Xylene 20:20 (v/v)
• System 2: Dioxane: Xylene: Petroleum ether 10:20:20 (v/v/v)
• System 3: n-Hexane: Ethyl acetate 98:2 (v/v)
For methanol extracts, the free metabolites were detected by TLC. The conjugated
◦
metabolites were liberated by heating the mixture with 2N HCl for 2 h at 100 C. After
cooling, the mixture was extracted with chloroform for three times, and the combined
chloroform was dried over anhydrous sodium sulfate, filtered off, evaporated under vacuum,
and identified by TLC analysis using the solvent systems used for identification of crude
oil.
Authentic samples were run alongside as references and spots were detected by
exposing to a UV source at 254 nm and made visible by subjecting the plates to I2 vapor
after a preliminary spray with PdCl2 solution.
Radioactivity Measurements
The radioactivity in the oil and methanol extracts was measured directly by LSC
(Packard Model TRI-CARB 2300 TR) in vials using a dioxane-based scintillation cocktail,
which was composed of dioxane (1 L), naphthalene (100 g), PPO—2,5-diphenyloxazole
(10 g), and POPOP—1,4-di[2-(5-phenyloxazolyl)]-benzene (0.25 g). Radioactivity in flow-
ers or cakes (100 mg) was determined by digestion using H2O2 and Solusol followed by
LSC. Silica gel plates were separately scraped (1-cm zones) and silica gel was eluted with
methanol followed by LSC. The counts were corrected for background and quenching
effect by the channel ratio method.
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