Residual Behavior of Malathion and its Metabolite Malaoxon in Four Varieties of Mango Fruits

Mango (Mangiferae indica) is one of the finest fruits and the most
important fruit crops in tropical and subtropical areas of the world. Increasing
commercial acreage and improved handling methods and shipping throughout
the world have increased the mango's popularity and availability in Europe
and US markets. Over the years, mango groves have spread to many parts of
the tropical and sub-tropical world, where the climate allows the mango to
grow best, and where most of the developing countries located. To date,
developing countries are facing massive economic and social problems. One
possible way out of this misery seems to be the opening of the economy in
order to participate in the gains arising from international trade. By increasing
export volume and export revenues, developing countries expect to create a
momentum and, thus, the impetus to stimulate the overall economy (Borchert,
2001).
Since Egypt located at mango production area, it is a big chance to
share in the international mango market by improving mango production
quantity and quality.
Mango suffers from several diseases at all stages of its life. All the
parts of the plant, namely, trunk, branches, twigs, leaf, petiole, flower and fruit
are attacked by a number of pests including insects. They cause huge
damage in quality and production of mango fruits (Ploetz, 2004 and Kaiser
and Saha, 2005).
Malathion, is a non-systemic insecticide. This insecticide is
cholinesterase enzyme inhibitor. Malathion can be bioactivated to malaoxon
via oxidation desulfuration by insect metabolism and then is transformed to
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isomalathion by thermal or photochemical isomerization. Isomalathion has
been identified in certain commercial formulations and is suspected to be a
prime agent in the death of 5 workers and the sickening of another 2800 in
Pakistan during a 1976 malaria-eradication program (Anping et al., 2013).
Malathion and malaoxon contain an asymmetric carbon atom, which leads to
the formation of two enantiomers, respectively. The isomerization of malathion
to isomalathion not only maintains the asymmetric carbon atom but also forms
a new asymmetric phosphorus atom, yielding four possible stereoisomers (Yu
et al., 2010). At present, malathion is still marketed and applied in its racemic
form despite the fact that the (R)-enantiomer shows a higher biological activity
than the (S)-isomer (Anping et al., 2013).
Malathion is effective in controlling many insects such as leaf eating
caterpillars, thrips, cockchafer larvae, cutworms, etc. in a range of crops
including vegetables, fruits, maize, sugar cane, sugar beet, tea, tobacco, and
ornamentals. However, malathion has, however, been reported to have
endocrine disrupting effects, Penalve et al. (2003). Malathion may have
harmful effects on large numbers of people are exposed to malathion in their
home or work environment, or through consumption of foods containing trace
levels of OP insecticides (Barr, 2004). Therefore, the environmental behavior
of malathion is increasingly being investigated.
This study is aiming to throw the light upon the residues of malathion and its
metabolite malaoxon on most famous mango varieties in Egypt as well as the
residues amount in the fruit with special reference to pre-harvest interval
(PHI).
MATERIAL AND METHODS
This study was carried out during 2011 and 2012 seasons on four
different mango varieties namely Alfouns, Zebdia, , Fajeri Klan , and Langra
(10 years old trees) grown in Elkatatba (Menoufia governorate).
Tested Pesticide:
Malathion was used as malatox 57% EC at the recommended dose (30
ml/liter of water) to control pests attacking mango trees. Malathion was
sprayed for one time.
a- Malathion diethy[(dimethoxythiophosphinothioyl)thio]butanedioate
b- Malaoxon (O- [1,2-bis(ethoxycarbonyl)ethyl] O,O-dimethyl
phosphorodithioate)
Fig 1: Chemical structure of malathion (a) and its isomer malaoxon (b)
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Sampling
Fruit samples of each mango variety were randomly picked up after one hour,
1, 3, 5 and 7 days after treatment.
Extraction:
The procedure of Lehotay et al. (2005) as a QuCHER (Quick, Cheap,
Effective and Rugged) method was used for extraction and purification of
pesticide residues from mango samples. The analysis procedure was done at
the Central Lab. of Residue Analysis of Pesticides and Heavy Metals in Food,
Agric. Res. Center, Egyptian Ministry of Agriculture. Fresh sample of 10 g was
weighed and mixed with 10 ml deionized water in a 50 ml PFTE tube by
shaking for one minute. Acetonitrile acidified with acetic acid (10 ml), 1.0 g
sodium acetate and 4.0 g anhydrous magnesium sulphate were added and
shaked vigorously for one minute. The samples were centrifuged at 4000 rcf
for 2 min. Six milliliters of the upper clear solution (extracts) were transferred
into 15 ml polyethylene tube contained 0.4 g primary secondary amine (PSA)
sorbent and 0.6 g anhydrous magnesium sulphate. The tubes were caped,
then the extract with the sorbent/ dessicant mixed vigorously for one minute
and centrifuged at 4000 rcf for 2 min. Four milliliters of the clear solution were
transferred into 15 ml glass tube and 50 μl tetradecan was added as keeper
and evaporated in turbovab at 40 oC to dryness. The residues were dissolved
in 2 ml of acetonitrile and then injected in GC.
For recovery studies, the samples were spiked with the studied
compounds before the corresponding extraction procedure has been done. A
representative 10 g portion of mango sample was weighed and fortified
homogeneously with appropriate volume of working standard solution and
followed the same previous procedure of determination.
Spiked levels were 0.03, 0.1 and 1.0 mg/kg. The obtained results were
corrected according to the recovery rate.
GLC procedures:
Assessment of malathion and malaoxon residues was carried out
according to the Official Methods of Analysis (Anonymous, 1995) using
Hewlett Packard gas liquid chromatography (HP 6890N) equipped with
nitrogen phosphorus detector (NPD), two columns, (HP PAS-5, NPD tested
Ultra 2 Silicone, 0.32 mm i.d., 0.52 μm film thickness and 25.0 m length and
HP PAS-1701, 0.32 μm i.d., 0.25 μm film thickness and 25 m length), HP
autosampler and HP computer under the following operating conditions:
Injector temperature = 225 oC, Detector temperature =280 oC, Flow rate of
nitrogen 60 ml/min (carrier + makeup), Column head pressure 80 kPa,
Splitless time 0.7 min.
The oven was programmed as follow:
Initial oven temperature: 90 oC, Initial oven time 2 min. using two ramps,
Ramp (1) Rate 20oC / min, Temp 150 oC, Time 0 min. and
Ramp (2) Rate 6oC / min, Temp 270 oC Time 15 min.
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The determined concentration in sample (Cs) (mg/kg) was calculated as
follows:
As / Ais Vf x Vtot
Cs = X Cst X
Ast / Aist Va x M
Where:
As = Peak area of analyte in sample
Ais = Peak area of ditalimiphos standard in sample
Ast = Peak area of analyte in standard run
Aist= Peak area of ditalimiphos standard in standard run
Cst = Concentration of standard (mg/L)
Vf = Final volume (ml)
Vtot.= Total extraction volume (ml)
Va = 40 ml
M = Sample weight in final volume (g)
Half life time (LT50) was calculated and pre-harvest interval (PHI) was
determined considering the MRL for malathion and its metabolite malaoxon
on mango which equal 0.2 and 0.05 mg/kg, respectively according to Annex II
Regulation of European Union (Anonymous, 2005).
RESULTS AND DISCUSSION
Pesticides residues in food stuff are one of the most limiting factors
affect the trade and export of all edible products. The objective of the present
investigation was monitoring the residues of malathion and its metabolite
malaoxon in four varieties of mango fruits through a period of time, and
predicting their PHI (Pre Harvest Interval). Since MRL (Maximum Residue
Limit) of malathion is 0.2 mg/kg, The estimated PHI (Pre Harvest Interval) was
found to be 3days. For malaoxon, MRL is 0.05 ppm and therefore, the
estimated PHI was found to be 2 days and the calculated LT50 values of
malation ranged from 0.96 to 2.8 days in these four varieties however, for
malaoxon it was ranged between 0.43 to 0.9 days (Table 1).These results are
in agreement with those reported by Lotfy et al. (2013) who found that LT50 on
zucchini was around 0.77 days and PHI was 0.5 days.
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Table 1. Malathion and malaoxon residues, dissipation % in different mango
varieties fruit after different intervals from treatment, half life time
(LT50), and pre-harvest interval (PHI)
Time
Postapplication
(days)
Mango Types
Al Founs Fager Kalan Langra Zebdia
Malathion
(mg/kg)
Malaoxn
(mg/kg
Malathion
(mg/kg)
Malaoxn
(mg/kg)
Malathion
(mg/kg)
Malaoxn
(mg/kg
Malathion
(mg/kg)
Malaoxn
(mg/kg)
0 0.49 0.15 2.3 0.35 1.7 0.13 0.87 0.18
1 0.25 0.07 1.5 0.06 0.61 0.05 0.46 0.07
3 0.13 ND 0.25 ND 0.1 ND 0.19 ND
5 0.04 ND 0.08 ND 0.06 ND 0.02 ND
7 ND ND ND ND ND ND ND ND
LT50 2.8 0.9 1.6 0.55 0.96 0.43 1.1 0.85
PHI 3 2 3 2 3 2 3 2
ND = Not Detected, LT50= Half Life Time
Malathion residues decreased with time and within every fixed time
interval, the decrease is a constant ratio from the amount already present at
the beginning of the interval, i.e., the rate of decrease in residues at any time
is directly proportional to the amount of the residues at that time. Rapid
disappearance of malathion and its metabolite malaoxon was observed in the
studied varieties of Egyptian mango (Fig. 2 and 3), with no residue levels
found after 5 days. These results are compatible with those of Mingjing et al.,
(2012). Control samples were fortified at the three levels of, 0.03, 0.1, and 1,
and average recovery percentages from spiked samples are listed in Table 2.
Table 2. Recovery percentage of malathion and malaoxon from mango at
three fortification levels
Malation Malaoxon
Fortification
level
Recovery%
0.03 ppm 0.1 ppm 1
ppm
88 91
97
0.03 ppm 0.1 ppm 1
ppm
85 90
99
It is clear from Table 2 that the recovery ranged from 88 to 97% for malathion
and 85 to 99% for malaoxon. The metabolites of malathion always were
proved to be more toxic than the parent compound (Zhang et al., 2013). The
higher toxicity of the metabolite is believed to be related higher bimolecular
rate constants of malaoxon with acetylcholinesterase and carboxylesterase.
Although malaoxon is a better inhibitor for carboxylesterase, malathion was
proved to be the best stable substrate for this enzyme. In vivo, however, the
inhibition reaction dominated the substrate reaction, resulting in the
metabolites being more toxic (Hassan and Dauterman, 1968). So, it was
important to determine the degradation rate and define the pre-harvest
intervals of both insecticides.
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0
0.5
1
1.5
2
2.5
0 1 2 3 4 5 6 7
Time (days)
Risidue (mg/kg)
Al Founs
Fager Kalan
Langra
Zebdia
Fig 2: Decay of malathion residues in four mango varieties
-0.05
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0 2 4 6 8
Time (days)
Residue (mg/kg)
Al Founs
Fager Kalan
Langra
Zebdia
Fig 3: Decay of malaoxon residues in four mango varieties
Recommendation
The safety period for the harvesting of different varieties of mango fruits in
Egypt should not before 3 days after treatment of the crop by malathion to
avoid the adverse toxic effect of malathion on human health.