Adverse effects of exposure to low doses of chlorpyrifos in lactating rats

Adverse effects of exposure to lowdoses of chlorpyrifos in lactating rats

Sameeh A Mansour and Abdel-Tawab H Mossa

AbstractThis study was conducted to shed light on the effect of exposure of lactating rat to chlorpyrifos (CPF). CPF wasorally administered to lactating rats at 0.01 mg kg�1 b.wt. (acceptable daily intake, ADI), 1.00 mg kg�1 b.wt. (noobserved adverse effects level, NOAEL) and 1.35 mg kg�1 b.wt. (1/100 LD50) from postnatal day 1 (PN1) untilday 20 (PN20) after delivery. Results indicated decreases in body weight and increases in relative liver andkidney weights of exposed dams. Significant damage to liver was observed via increased plasma levels ofaminotransferases (aspartate aminotransferase (AST) and alanine aminotransferase (ALT)) lactatedehydrogenase (LDH) and g-glutamyle transferase (g-GT) in a dose-dependent manner. At two high dosesof CPF (1.00 and 1.35 mg kg�1 b.wt.), the lactating mothers showed significant decrease in the activity of cho-linesterase (ChE). Lipid peroxidation was significantly increased, while glutathione s-transferase (GST) andsuperoxide dismutase (SOD) were significantly decreased compared to control. At high dose of CPF (1.35mg kg�1 b.wt.), total protein and uric acid levels were significantly increased. CPF caused dose-related histo-pathological changes in liver and kidney of the CPF-treated dams.

KeywordsChlorpyrifos, lactation rats, lipid peroxidation, oxidative damage, liver and kidneys dysfunction

Introduction

Chlorpyrifos, CPF (0,0-diethyl-0-3,5,6-trichloro-2-

pyridyl phosphorothioate), is a broad-spectrum,

chlorinated organophosphate (OP) insecticide, con-

sidered as one of the largest selling OP in the world,

and has both agricultural and urban uses. Some esti-

mates for pregnant women and children indicate that

CPF exposures exceed no observable adverse effect

level (NOAEL), even in scenarios of common use

(Davis and Ahmed, 1998; Gurunathan et al., 1998).

Mothers may be exposed to lipophilic chemical (e.g.

CPF) from various sources including air, food, water

and occupational and household environments.

Lipophilic chemicals can be stored and accumulated

over time in body fat and can then be mobilized into

milk during lactation. Generally, chemicals enter

breast milk by passive transfer from plasma, and their

concentration in milk is proportional to their solubi-

lity and lipophilicity (Anderson and Wolff, 2000),

showing the relevance of the present issue with

human health concerns. Previous study reported that

OP pesticides (e.g. CPF) can be accumulated and

excreted in human milk (Marty et al., 2007; Mattsson

et al., 2000; Salas et al., 2003; Sanghi et al., 2003).

Also, it has been reported that milk CPF concentra-

tions were up to 200 times those in blood of pregnant

rats dosed with CPF (Mattsson et al., 2000). The latter

authors also made an exposure estimate of CPF to

nursing pups and reported it as roughly 0.1 mg kg�1

b.wt. via nursing of dams exposed by gavage to

5 mg kg�1 b.wt. The nursing dose to pups was attenu-

ated greatly from gavage dose to their dams.

In the context of chemical toxicology, it is important

to identify the chemical responsible for human

toxicity. Biological monitoring is a well-established

Environmental Toxicology Research Unit (ETRU), PesticideChemistry Department, National Research Centre (NRC),Cairo, Egypt

Corresponding author:Abdel-Tawab H Mossa, Environmental Toxicology Research Unit(ETRU), Pesticide Chemistry Department, National ResearchCentre (NRC), Tahrir Str, Dokki, Cairo 12311, EgyptEmail: [email protected]

Toxicology and Industrial Health27(3) 213–224ª The Author(s) 2011Reprints and permission:sagepub.co.uk/journalsPermissions.navDOI: 10.1177/0748233710384054tih.sagepub.com

technique for assessing intakes and uptakes of

toxic chemicals following either occupational or

environmental exposure. For assessing human

exposure to many of the OP insecticides and for linking

OP exposures in pregnant women to subsequent

adverse birth outcomes, the metabolite containing the

organic moiety, such as TCPy in the case of CPF, has

been used as a more specific urinary biomarker

(Berkowitz et al., 2004; Eskenazi et al., 2004; Nolan

et al., 1984).

In fact, the toxicity of many xenobiotics is associ-

ated with the production of oxygen-free radicals,

more generally known as ‘‘reactive oxygen species’’

(ROS), which are not only toxic themselves but are

also implicated in the pathophysiology of many dis-

eases (Abdollahi et al., 2004; Akhgari et al., 2003).

The harmful effects of ROS are balanced by the

antioxidant action of nonenzymatic antioxidants in

addition to antioxidant enzymes (Halliwell, 1996).

Despite the presence of the cell’s antioxidant defence

system to counteract oxidative damage from ROS,

oxidative damage accumulation during the life cycle

has been proposed to play a key role in the develop-

ment of age-dependent diseases such as cancer, arter-

iosclerosis, arthritis, neurodegenerative disorders and

other conditions (Halliwell and Gutteridge, 1999).

It has been reported that OP insecticides may induce

oxidative stress following acute exposure in humans

(Banerjee et al., 1999) and animals (Mansour and

Mossa, 2009, 2010a).

Breastfeeding is one of the most important contri-

butors to infant health and prolonged breastfeeding

protects babies from common childhood infections

through mechanisms that are interactive, adaptive and

extend into childhood (Kramer and Kakuma, 2002).

Also, breastfeeding offers a range of health benefits

for mothers, including a reduced risk of ovarian

(Rosenblatt and Thomas, 1993) and pre-menopausal

breast cancer (Collaborative Group on Hormonal

Factors in Breast Cancer, 2002). However, potential

risks associated with breast-feeding need to be

factored into the overall public health assessment

when mothers are encouraged to breast-feed their new-

born infants (Gallenberg and Vodicnik, 1989). This

could be seen as a logical demand if we considered that

lactation may pose a ‘sort of stress’ to the mothers.

Thus, such mothers will be more vulnerable, than non-

lactating mothers, to other chemical stressors. To the

best of our knowledge, toxicological information of

pesticides on lactating animals (e.g. rats) are very lim-

ited. So, the present study was undertaken on lactating

rats to evaluate the effect of exposure to CPF at low

doses. Oxidative stress, lipid peroxidation (LPO), as

well as liver and kidney dysfunction will be the criteria

of assessing the exposure effects.

Materials and methods

Chemicals

Chlorpyrifos ‘CPF’ (M.Wt. 350.6; 99% purity) was

obtained from Dow AgroSciences (Indianapolis,

Indiana, USA) and 2-thiobarbituric acid (TBA; 2,

6-dihydroxypyrimidine-2-thiol; TBA) was purchased

from Merck (Germany). All other chemicals were of

reagent grade and were obtained from the local

scientific distributors in Egypt. The kit of lactate

dehydrogenase (LDH) was obtained from Spinreact

(Santa Coloma, Spain), gamma-glutamyl transferase

(g-GT) from Greiner Diagnostic GmbH (Bahlingen,

Germany), protein from Stanbio Laboratory (Texas,

USA) and albumin from Biogamma (Roma, Italy). The

kits of superoxide dismutase (SOD), glutathione s-

transferase (GST), aminotransferases (ALT and AST),

uric acid and creatinine were obtained from Biodignos-

tic, and cholinesterase (ChE) from Diamond Diagnostic

(Egypt).

Animals and housing

The healthy male and female albino rats of the Wistar

strain Rattus norvegicus, weighing 200–220 g, were

obtained from the Animal Breeding House of the

National Research Centre (NRC), Dokki, Cairo,

Egypt. Rats were allowed to acclimate to laboratory

conditions for at least 1 week before breeding. Thirty

virgin female rats were distributed into 10 cages. In

each cage, one male was placed for overnight and the

presence of spermatozoa was checked in the vaginal

smear the following morning. This day was connoted

as gestation day 0 (GD 0). At this time, pregnant

females were individually housed in clean plastic

cages in the laboratory animal room (23�C + 2�C)

on the standard pellet diet and tap water ad-libitum,

a minimum relative humidity of 40% and a 12 h

dark/light cycle. The day of parturition, was consid-

ered day 0 of lactation, postnatal day 0 (PND 0).

Offspring of each litter were randomly reduced to 8

pups of the equal number of sex’s +1, it has been

shown that this procedure maximizes the lactation

performance (Fishbeck and Rasmussen, 1987).

The experimental work on rats was performed with

the approval of the Animal Care & Experimental

214 Toxicology and Industrial Health 27(3)

Committee, National Research Centre, Cairo, Egypt,

and according to the guidance for care and use of

laboratory animals (NRC, 1996).

Experimental design

On the first day after parturition (PND1), 20 individu-

ally housed dams were segregated into four different

groups, five each. CPF was dissolved in corn oil and

administered by gavage at a volume of 0.5 mL/rat.

Three groups of dams were given daily, via oral route,

doses equaled to 0.01 mg kg�1 b.wt. (ADI), 1.00 mg

kg�1 b.wt. (NOAEL), and 1.35 mg kg�1 b.wt. (1/100

LD50), of CPF according to Tomlin (2005), during the

lactation period (PND 1 to PND 20), respectively. The

fourth group of dams was used as control and received

the equivalent volume of corn oil. Dosages of CPF

administrated were adjusted daily for body weight

changes and given at approximately the same time

each morning. The animal’s cages were cleaned daily

to minimize potential contamination.

Blood collection, body weight andorgan weight ratio

Dam’s body weight was recorded daily prior to dosing.

The blood samples were drawn from all dams on

postnatal day 21 (PND 21) under ether anesthesia by

puncturing the retero-orbital venous plexus of the ani-

mals with a fine sterilized glass capillary and collected

into heparinized tubes. Within 30 min of blood collec-

tion, the plasma samples were drawn from blood after

centrifugation at 3500 rpm (600g) for 10 min at 4�C,

using Hereaeus Labofuge 400R, Kendro Laboratory

Products GmbH, Germany, to separate the plasma. The

plasma was kept in a deep freezer (�20�C) until

analyzed within 10 days maximum.

After blood collection, the dams were sacrificed by

cervical dislocation. Liver and kidneys of dams were

quickly removed and weighted individually. Then, the

relative organs weights to the body weights were

calculated.

Biochemical analyses

Antioxidant enzyme assays. SOD and GST were

determined in plasma according to the manufacturer’s

instructions referred to in Woolliams et al. (1983) and

Habig et al. (1974), respectively. The activities were

expressed in terms of mmol/min/mg protein for both

enzymes.

Estimation of lipid peroxidation. Malondialdehyde

(MDA), as a marker for LPO, was determined by the

double-heating method of Draper and Hadley (1990).

The principle of the method is based on spectrophoto-

metric measurement of the color produced during the

reaction of TBA with MDA. For this purpose, 2.5 mL

of 100 gl�1 trichloroacetic acid (TCA) solution was

added into 0.5 mL plasma in a centrifuge tube and

placed in a boiling water bath for 15 min. After

cooling under tap water, the mixture was centrifuged

at 600g for 10 min, and 2 mL of the supernatant was

transferred into a test tube containing 1 mL of 6.7 gl�1

TBA solutions and placed again in a boiling water

bath for 15 min. The solution was then cooled under

tap water and its absorbance was measured spectro-

photometrically at 532 nm. The concentration of

MDA was calculated by the absorbance coefficient

of MDA-TBA complex 1.56 � 105 cm-1 M-1 and

expressed in nmol/mL.

Liver and kidneys markers. The measurement of

plasma cellular enzymes such as aminotransferases

(AST; EC 2.6.1.1 and ALT; EC 2.6.1.2), lactate

dehydrogenase (LDH; EC 1.1.1.27) and g-glutamyl-

transferase (GGT) were determined by the methods

of Reitman and Frankel (1957), Tietz (1995) and

Szasz (1969), respectively. The activity of AST, ALT,

LDH and GGT were expressed in terms of U/L. The

activity of plasma cholinesterase (BChE;

EC 3.1.1.8) was determined by the methods of Ellman

et al. (1961) and expressed as U/mL. The concentra-

tion of albumin (g/dL), total protein (g/dL), uric acid

and creatinine (mg/dL) were determined by the meth-

ods of Westgard and Poquette (1972), Gornal et al.

(1949), Barham and Trinder (1972) and Bartels

et al. (1972), respectively.

Histopathological studies

Liver and kidney samples were dissected and fixed in

10% neutral formalin, dehydrated in ascending grades

of alcohol and imbedded in paraffin wax. Paraffin

sections (5 mm thick) were stained for routine histolo-

gical study using haematoxylin and eosin (H&E). For

each rat, two slides were prepared; each slide

contained two sections for each organ. Ten field areas

for each section were selected and examined for

histopathological changes (64�) under light micro-

scope. The liver fields were scored as follows: nil

(normal appearance) ¼ 0%, þ ¼ mild (cellular disrup-

tion in less than 20% of field area), þþ ¼ moderate

Mansour and Mossa 215

(cellular disruption of 20% to less than 40% of field

area) and þþþ ¼ severe (cell disruption of 40 to less

than 70% of field area). The kidney fields were scored

based on tubular injury as described above. Such

quantitative assessment of histopathological injury has

been performed by previous investigators (Kerem

et al., 2007).

Spectrophotometric measurements

The spectrophotometric measurements were performed

by using a Shimadzu UV-VIS Recording 2401 PC

(Japan).

Statistics

The data were analyzed by using SPSS (version 14.0)

for Windows and expressed as means + SD. Paired

samples t test was used to compare between the data

of the control and those of treatments.

Results

Body and relative organs weights

There was no significant difference in body and liver

and kidney weights in lactating mothers exposed to

0.01 mg kg�1 b.wt. of CPF at any time point in the

study, compared to control groups (Figure 1). The

treated groups with 1.00 mg kg�1 b.wt. and 1.35 mg

kg�1 b.wt. of CPF had a significant decrease in body

compared to the corresponding control group

(Figure 1A). The significant decrease in body weight

was observed from the PND 10 to the end of experi-

ment (PND 21). The reduction in body weight, if

calculated, accounted to �0.98%, �16.35% and

�18.53% on PND 21 with respect to control values,

for 0.01, 1.00 and 1.35 mg kg�1 b.wt. groups, respec-

tively. A dose-dependent decrease in body weight of

mothers exposed to CPF was observed. In contrast,

there was a significant increase in relative liver

weight in mothers exposed to1.00 and 1.35 mg kg�1

b.wt. of CPF and significant increase in relative

kidney weight in mothers exposed to 1.35 mg kg�1

b.wt. of CPF (Figure 1B).

Oxidative stress markers

Results of oxidative stress markers are shown in

Table 1. There was no significant difference in SOD

and GST activity and LPO level in mothers exposed

to 0.01 mg kg�1 b.wt. of CPF. But there were signif-

icant decreases in the activity of SOD and GST and

significant increase in LPO level in mothers exposed

to either 1.00 or 1.35 mg kg�1 b.wt. of CPF compared

to control groups in a dose-dependent manner. The

changes in the activity of SOD and GST (if calculated)

accounted to �5.59%, �13.14% and �19.01% and

�16.0%,�46.0% and�51.33% with respect to control,

for 0.01, 1.00 and 1.35 mg kg�1 b.wt. of CPF,

respectively.

Liver and kidneys markers

Treatment of lactating mothers with CPF caused

changes in the levels of AST, ALT, LDH, GGT and

ChE compared to control groups in a dose-

dependent manner (Table 2). A significant increase

(p � 0.05) in the activity of AST was observed in the

treatment with 0.01 mg kg�1 b.wt. At 1.0 mg kg�1

b.wt. CPF, except ALT, the other major parameters

recorded significant elevation compared to the corre-

sponding control values. At high dose of CPF (1.35

mg kg�1 b.wt.), the lactating mothers showed signif-

icant increase in the activity of AST, LDH, GGT

(p � 0.01) and ALT (p � 0.05). AST/ALT value

accounted to 1.13 for the control group and 1.51,

1.63 and 1.66 for the three tested doses of CPF,

respectively. At two high doses of CPF (1.00 and

1.35 mg kg�1 b.wt.), the lactating mothers showed

significant decrease in the activity of ChE and the

change in ChE activity for the CPF groups, if

calculated, accounted to �11.11%, �15.74% and

�25.46% for 0.01, 1.00 and 1.35 mg kg�1 b.wt. of

CPF, respectively (Table 2). As shown in Table 3, the

high dose of CPF (1.35 mg kg�1 b.wt.) caused signif-

icant increase in total protein, globulin and uric acid

(p � 0.05) and decrease in albumin (p � 0.01) con-

centrations in the plasma of the tested mothers. Crea-

tinine levels in exposed mothers did not show

significant alteration due to CPF-exposure at any

doses.

Histopathology changes

Histopathological findings of liver and kidney for

various treatment groups are presented in Table 4.

In light microscopic examinations, histopathologi-

cal changes were observed in liver and kidney of

all exposed groups compared to control ones. How-

ever, these changes were more frequent in CPF-

treated groups with 1.00 mg kg�1 b.wt. and 1.35

mg kg�1 b.wt. The liver and kidney sections of

control animals showed normal structures. With

respect to the hepatic histoarchitecture of the


CPF-treated animals, dilatation in central vein,

degeneration in hepatocytes, congestion of portal

vein and proliferation in bile duct, focal necrotic and

fibrosis were the main findings in CPF-treated

groups. In cases of kidney, the renal histoarchitec-

ture of the CPF-treated animals, degeneration and

necrosis in the epithelial cells lining the tubules

and proliferation of the endothelial cells lining the

tuft glomerulus were the main findings in CPF-

treated group.

Discussion

In the present study, oral administration of CPF to

lactating mothers resulted in significant decrease, in

a dose-dependent manner, in body weight and signif-

icant increase in relative liver and kidney weights.

Reduction in body weight observed in CPF-treated

dams may be a result of the combination of increased

degradation of lipids and proteins as a result of

the direct effects of CPF as an organophosphate

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20151051

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Figure 1. Body weight (A) and relative organs weights (B) of lactating mothers exposed to chlorpyrifos (CPF) duringlactation period (postnatal day 1 [PND1] to PND 20). Each value is a mean + SD; statistical difference from the control:* significant at p � 0.05 and ** highly significant at p � 0.01. n ¼ 5 animals per group.


compound (Goel et al., 2005), as well as reduction of

food intake by lactating mothers (untabulated data),

which may cause a negative energy balance. Reduc-

tion of body weight and feed consumption in lactating

dams following oral exposure to CPF were previously

reported by Maurissen et al. (2000). In this respect, dur-

ing lactation, in many human mothers (Butte et al.,

1984) and animals (McNamara and Hillers, 1986),

decrease of the body weight and percentage body fat

is seen, presumably because the energy needs of

lactation plus normal energy needs exceed their energy

intake. The increase in relative liver weight is attributed

to the increase of UDP-glucuronyltransferase (UDPGT)

enzyme activity, which elevates in liver, brain and kid-

ney of rats orally administered some OPs (Bulusu and

Chakravarty, 1986; Cook et al., 1997; Mahboob and

Siddiqui, 2001). The present findings related to changes

in body and internal organs weights were reported by

other investigators (Ambali et al., 2007; Mansour

et al., 2008; Mansour and Mossa, 2009). In previous

Table 1. LPO level, SOD and GST enzymes activity in plasma of lactating mothers exposed to CPF during the sucklingperiod (PND1 to PND 20)a

Dose (mg kg-1 b.wt.) LPO (nmoL/mL) GST (mmoL/mg protein) SOD (mmoL/mg protein)

0.00 1.43 + 0.08 1.50 + 0.12 120.51 + 7.140.01 1.47 + 0.10 1.26 + 0.16 114.13 + 10.111.00 1.66 + 0.12b 0.81 + 0.19c 105.51 + 5.22b

1.35 1.77 + 0.15b 0.73 + 0.02c 98.81 + 6.08c

Abbreviations: LPO: lipid peroxidation, SOD: superoxide dismutase, GST: glutathione s-transferase, CPF: chlorpyrifos, PND: postnatalday.a Each value is a mean + SD. n ¼ 5 animals per group.b Statistical difference from the control: significant at p� 0.05.c Statistical difference from the control: highly significant at p � 0.01.

Table 2. Plasma AST, ALT, LDH, GGT and ChE enzymes activity in lactating mothers exposed to CPF during the sucklingperiod (PND1 to PND 20)a

Dose (mg kg-1 b.wt.) AST (U/L) ALT (U/L) AST/ALT value LDH (U/L) GGT (U/L) ChE (U/mL)

0.00 43.44 + 2.89 38.37 + 1.25 1.13 + 0.04 173.14 + 5.69 10.06 + 1.28 2.16 + 0.120.01 55.00 + 3.96b 36.36 + 1.12 1.51 + 0.06b 183.50 + 20.81 11.36 + 2.76 1.92 + 0.181.00 62.23 + 4.31c 38.07 + 2.24 1.63 + 0.02b 231.26 + 20.84b 23.04 + 4.21c 1.82 + 0.09b

1.35 76.51 + 4.67c 46.19 + 3.45b 1.66 + 0.02c 255.02 + 22.09c 25.09 + 3.89c 1.61 + 0.07c

Abbreviations: AST: aspartate aminotransferase, ALT: alanine aminotransferase, LDH: lactate dehydrogenase, GGT: g-glutamyl trans-ferase, ChE: Cholinesterase, CPF: chlorpyrifos.a Each value is a mean + SD; n ¼ 5 animals per group.b Statistical difference from the control: significant at p � 0.05.c Statistical difference from the control: highly significant at p � 0.01.

Table 3. Plasma total protein, albumin, globulin, A/G ratio, uric acid and creatinine level in lactating mothers exposed toCPF during the suckling period (PND1 to PND 20)a

Dose (mg kg-1

b.wt.)Total protein

(g/dL)Albumin(g/dL)

Globulin(g/dL)

Albumin/globulin(A/G) ratio

Uric acid(mg/dL)

Creatinine(mg/dL)

0.00 7.10 + 0.68 3.40 + 0.13 3.70 + 0.32 0.92 + 0.04 3.99 + 0.41 0.61 + 0.100.01 7.19 + 0.71 3.29 + 0.14 3.90 + 0.45 0.84 + 0.06 3.84 + 0.56 0.65 + 0.131.00 8.22 + 1.19 3.15 + 0.29 5.07 + 0.61b 0.62 + 0.02b 5.12 + 0.39b 0.65 + 0.121.35 9.17 + 1.03b 2.85 + 0.10c 6.32 + 0.11b 0.45 + 0.01c 5.45 + 0.89b 0.81 + 0.14

Abbreviations: A/G: albumin/globulin, CPF: chlorpyrifos.a Each value is a mean + SD; n ¼ 5 animals per group.b Statistical difference from the control: significant at p � 0.05.c Statistical difference from the control: highly significant at p � 0.01.


studies, administration of CPF at 1.0 mg kg�1 b.wt. did

not induce significant changes in body weight gain,

either in dams (Maurissen et al., 2000) or female rats

(Yano et al., 2000).

In this context, the maximum-tolerated-doses

(MTD) was initially based on a weight gain decre-

ment observed in the subchronic study; i.e., the high-

est dose that caused no more than a 10% weight gain

decrement (Interagency Staff Group on Carcinogens,

1986). It is likely to state that the decrease in body

weight gain may reach up to 20% in toxicology

studies designated for risk assessment at MTD of a

toxicant. The decrease in body weight gain in the

present study at doses 1.0 and 1.35 mg kg�1 b.wt.

were far of MTD for CPF, which was administrated

in corn oil. It is documented that gavage in corn oil

enhances absorption of CPF across the gastrointest-

inal tract (GI) mucosa, thus increasing the pesticide

toxicity (Conolly et al., 1999). On the other hand,

Marty et al. (2007) reported that dietary CPF treat-

ment at 5 mg kg�1 b.wt. in lactating rats had blood

CPF levels that were some 13� lower than the same

dose by oral gavage. For this reason, Conolly et al.

(1999) stressed cautions against use of corn oil gavage

administration in risk assessment of organic com-

pounds, generally. However, we cannot ignore the

effect of exposure to a toxicant in conjunction with

fats in the food.

Some studies have shown that oxidative stress

could be an important component for the mechanism

of toxicity of organophosphate insecticides (OPIs).

OPIs may induce oxidative stress leading to genera-

tion of free radicals and alteration in antioxidants or

ROS scavenging enzymes (Banerjee et al., 1999;

Mansour and Mossa, 2009, 2010a; Ranjbar et al.,

2002). Therefore, OPI-may enhance lipid peroxida-

tion (LPO) by directly interacting with the cellular

membrane and ROS. Previous studies have suggested

LPO as one of the molecular mechanisms involved in

OPIs-induced toxicity (Bagchi et al., 1995; Mansour

and Mossa, 2010a).

Plasma lipid peroxidation was estimated in the

form of thiobarbituric acid reactive substances

(TBARS) produced. Significant increase in TBARS

was observed in mothers exposed to 1.00 and 1.35

mg kg�1 b.wt. of CPF when compared with control

(Table 1). Increased formation of peroxide (TBARS)

could be due to the increased peroxidation of

membrane. Literature shows the basic process by

which any and related radicals such as superoxide and

hydroxy radicals cause membrane damages in LPO

(Bachowski et al., 1997; Bagchi et al., 1995; Mansour

and Mossa, 2010a; Yamano and Morita, 1992). Our

results revealed that CPF caused a statistically signif-

icant decrease in the activity of SOD and GST and

significant increase in LPO level of mothers exposed

to CPF. The decrease in the activity of SOD in CPF-

intoxicated animals may be due to the consumption of

this enzyme in converting the O2- to H2O, which cat-

alyzed by SOD. Considering that GST are detoxifying

enzymes that catalyze the conjugation of a variety of

electrophilic substrates to the thiol group of GSH,

producing less toxic forms (Hayes and Pulford,

1995), the significant decrease of GST activity may

indicate insufficient detoxification of CPF in rats.

However, the activation of CPF to corresponding

oxons occurs through oxidative desulphuration

mediated by the cytochrom P450. CPF induces

depression in the activity of both nicotinamide ade-

nine dinucleotide phosphate (NADPH) cytochrome-

c-reductase and nicotinamide adenine dinucleotide

(NADH) cytochrome-c-reductase in intoxicated rats

(Goel et al., 2007). It was suggested that the sulphur

released during desulfuration of an OP insecticide

Table 4. Histopathological changes in the liver and kidneys of lactating-mothers exposed to CPF during the sucklingperiod (PND1 to PND 20), based on scoring severity of injurya in both organs

Dose (mgkg�1 b.wt.)

Hepatic injury Renal injury

Observation Severity Observation Severity

0.00 Normal Nil Normal Nil0.01 Congestion; degeneration; hyperplasia of bile duct þ Degeneration þ1.00 Congestion; necrosis; inflammatory cell infiltration in

portal area; fibrosis; hyperplasia of bile ductþþ Glomerulus hyperplasia;

necrosis renal tubulesþþ

1.35 Congestion; necrosis þþ Hyperplasia in glomerulustubules; degeneration

þþ

Abbreviation: CPF: chlorpyrifos.a Scores in terms of numerical values are mentioned in histopathological studies section, number of slides ¼ 10/group/organ.


may bind not only to the cytochrom P450 but also to

NADH cytochrome-c-reductase, culminating in the

overall reduced activity of this monooxygenase

(Kamataki and Neal, 1976). Alternatively, NADPH

cytochrome-c-reductase is also involved in initiating

the process of NADPH-dependent lipid peroxidation

in the microsomal membranes, and increased lipid per-

oxidation as a result of CPF intoxication may also be

the cause of depressed enzymatic activity (Sevanian

et al., 1990). These indirectly suggest an increased

production of oxygen-free radicals in rats. Highly

reactive oxygen metabolites, especially hydroxyl

radicals, act on unsaturated fatty acids of phospholipid

components of membranes to produce MDA, an LPO

product (Mansour and Mossa, 2009, 2010a).

ROS damage cells by free radical transfer from an

initial metabolic by-product to a variety of critical

biomolecules including DNA, proteins and lipids.

Free radical addition increases the likelihood of

modification through conjugation with electrophiles

or covalent bond breakage. Physical damage, mutation,

or loss of function in the short term can cause hepato-

cyte death and over the longer term can contribute to

the pathophysiology of liver disease (Murray et al.,

2006). This is in addition to the cellular stress and

energy consumption caused by redox cycling, which

on its own can overwhelm the antioxidant capacity of

hepatocytes and result in cell death. A major pathophy-

siologic concept that has emerged is that oxidative

stress contributes to many diseases, including cancer,

cardiovascular disease, liver cirrhosis, diabetes and

others related to aging (Murray et al., 2006).

Liver is the main tissue in detoxification and

metabolism of chemicals and along with kidney faces

the threat of maximum exposure to xenobiotics and

their metabolic by-products. This may impair its

regular function due to xenobiotic modification in

detoxification processes. In fact, the susceptibility of

liver and kidney tissues to this stress due to exposure

to pesticides is a function of the overall balance

between the degree of oxidative stress and the antiox-

idant capability (Khan et al., 2005). However, several

of soluble enzymes of blood plasma have been consid-

ered as indicators of hepatic dysfunction and damage.

LDH is one of the metabolic requirements of tissue and

involved in energy production. LDH activity indicates

the switching over of anaerobic glycolysis to aerobic

respiration. It can be used as an indicator for cellular

damage, clinical practice and cytotoxicity of toxic

agents (Bagchi et al., 1995). Also, g-glutamyl

transferase catalyzes the transfer of g-glutamyl group

from a g-glutamyl peptide to an amino acid or another

peptide. This enzyme is widely used as a biomarker in

preneoplastic lesions of the liver during chemical

carcinogenesis (Peraino et al., 1983). Our results

demonstrated that CPF may affect liver metabolism

and the leakage of certain intracellular enzymes,

suggesting damage in hepatocytes. Since AST/ALT

leaking to plasma is very sensitive marker of hepato-

cyte injury, increase in plasma enzymes may occur due

to minor liver injury in response to chemicals with low

histological changes (Williams and Iatropoulos,

2002). In the present study, the increment of the

activities of AST, ALT, LDH and GGT in plasma

could be attributed to the leakage of these enzymes

from the liver cytosol into the blood stream (Navarro

et al., 1993), which indicated liver damage and disrup-

tion of normal liver function (Shakoori et al., 1994; de

Boer et al., 2000; Kuester et al., 2002; El Sakka et al.,

2002). Also, AST/ALT values were higher than con-

trol of exposed mothers and ranged ‘between 1.51 to

1.66,’ which seemed to be higher than normal value

(1.00-1.50; Caglar and Kolankaya, 2007).

Cholinesterase (ChE, EC 3.1.1.8), also known as

pseudocholinesterase, has been recognized as an

enzyme that hydrolyzes choline esters. It has already

been mentioned that ChE is synthesized mainly in

hepatocytes and secreted into the blood stream

(Brown et al., 1981). ChE activity is reduced in liver

dysfunction due to reduced synthesis, in contrast to

other serum enzymes associated with the clinical

assessment of liver function whose activities increase

as a result of increased release from their cellular

sources following cell membrane damage (Moss and

Henderson, 1999). In this respect, changes in ChE

activity reflect the changes in hepatocellular functions

and have been regarded as sensitive indicators of the

diminished synthetic capacity of the hepatic parench-

yma (Adolph, 1979). In the present study, plasma ChE

activity of dams given 1.00 and 1.35 mg kg-1 b.wt. of

CPF was significantly decreased (15.74 and 25.46%)

compared to the control.

In previous studies, CPF was administered to rat

dams by gavage in corn oil (Maurissen et al., 2000)

and to female rats in feed (Yano et al., 2000). In both

studies, the administered dose of CPF was 1.0 mg

kg�1 b.wt., resembling the median dose tested in the

present study. Among comparable biochemical

criteria measured in these studies, the inhibition of

ChE, which was accounted to 31.1% and 81.0%,

respectively, compared to 15.74% in the present inves-

tigation. It is known that the toxicity of a toxicant may


be affected by several factors, such as strain of rats,

route of administration, dose and lactation statutes.

Total protein and A/G ratio is done as a routine test

to evaluate the toxicological nature of various chemi-

cals. Increases of total protein and decrease of A/G

ratio were observed in the present study following

CPF treatment. In relation to decrease in the A/G

ratio, the albumin level was also decreased, suggest-

ing high plasma globulin levels reflecting high protein

in CPF-treated mothers. The increase of total protein

in CPF-treated groups may be due to (a) due to

production of enzymes lost as a result of tissue

necrosis (b) to meet increased demand detoxifying the

pesticide might necessitate enhanced synthesis of

enzyme proteins (Gill et al., 1990, 1991), and (c) may

be due to kidney dysfunction (Mansour and Mossa,

2005; Mansour and Mossa, 2010a).

Kidney has a relatively high level of enzyme

activities, and the role of this organ in converting

xenobiotics and endogenous substances into excretory

forms is considerable (De Kanter et al., 2002). In the

present study, exposed mothers to CPF at 1.00 mg g�1

b.wt and 1.35 mg g�1 b.wt. showed significant

increases in uric acid concentration in the plasma

compared to the control, while creatinine levels did

not show significant alteration. In fact, uric acid is the

end product of the catabolism of tissue nucleic acid,

i.e., purine and pyrimidine based metabolism, and the

increase in uric acid may be due to degradation of

purines and pyrimidines or to an increase of uric acid

level by either overproduction or inability of excretion

(Mansour and Mossa, 2005).

The histopathological lesions observed in the liver

and kidneys of CPF-treated animals are in corrobora-

tion with the observed biochemical changes. These

observations indicated marked changes in the overall

histoarchitecture of liver and kidney in response to

CPF, which could be due to its toxic effects primarily

by the generation of ROS, causing damage to the

various membrane components of the cell. Our results

are supported by other studies conducted on CPF and

other OP insecticides (Mansour et al., 2008; Mansour

and Mossa, 2010a, b; Wang and Zhai, 1988).

Conclusion

Exposure of lactating dams to low dose levels of the

OP insecticide, CPF, induced pronounced alterations

in LPO, oxidative damage, and liver and kidneys

dysfunctions. Compared to results of other studies

(Maurissen et al., 2000; Yano et al., 2000) conducted

at different dose levels of CPF, including 1.0 mg kg�1

b.wt., the observed differences in toxicity results

would be attributed to a number of factors, such as

strain of rats, route of administration, exposure

period, dose and lactation statutes. It has been

reported that CPF exposures exceed no observable

adverse effect level (NOAEL) for pregnant women

and children, even in scenarios of common use (Davis

and Ahmed, 1998; Gurunathan et al., 1998). This lead

to paying more attention to the risks posed to lactating

women especially in developing countries, where

women are usually involved in agricultural practices.

Moreover, lactation transfers of chlorinated OPIs such

as CPF cause detrimental hazards to breast-feeding

infants (Mansour and Mossa, 2010b).

Acknowledgement

The authors are grateful to Prof. Dr Adel Mohamed Bakeer

Kholoussy, Professor of Pathology, Faculty of Veterinary

Medicine, Cairo University, for reading the histopathologi-

cal sections.

Funding

The author(s) received no financial support for the research

and/or authorship of this article.

References

Abdollahi M, Ranjbar A, Shadnia S, Nikfar S, and Rezaie

A (2004) Pesticides and oxidative stress: a review.

Medical Science Monitor 10(6): RA141–RA147.

Adolph L (1979) Diagnostische bedeutung der cholinesterase-

bestimmung im menschlichen serum. Med Wschr 121:

1527–1530.

Akhgari M, Abdollahi M, Kebryaeezadeh A, Hosseini R,

and Sabzevari O (2003) Biochemical evidence for free

radical induced lipid peroxidation as a mechanism

for subchronic toxicity of malathion in blood and

liver of rats. Human and Experimental Toxicology

22: 205–211.

Ambali S, Akanbi D, Igbokwe N, and Shittu M (2007)

Evaluation of subchronic chlorpyrifos poisoning on

hematological and serum biochemical changes in mice

and protective effects of vitamin C. Toxicology Sciences

32: 111–120.

Anderson HA, Wolff MS (2000) Environmental contami-

nations in human milk. Journal of Exposure Analysis

and Environmental Epidemiology 10: 755–760.

Bachowski S, Xu Y, Stevenson DE, Walborg EF, and

Klaunig JE (1997) Role of oxidative stress in the

mechanism of dieldrin’s heptotoxicity. Annals of

Clinical Laboratory Science 27: 196–209.


Bagchi D, Bagchi M, Hassoun EA, and Stohs SJ (1995) In

vitro and in vivo generation of reactive oxygen species,

DNA damage and lactate dehydrogenase leakage by

selected pesticides. Toxicology 104: 129–140.

Banerjee BD, Seth V, Bhattacharya A, Pasha ST, and

Chakraborty AK (1999) Biochemical effects of pesti-

cides on lipid peroxidation and free radicals scavengers.

Toxicology Letter 107: 33–47.

Barham D, Trinder P (1972) An improved colour reagent

for the determination of blood glucose by the oxidase

system. Analyst 97: 142–145.

Bartels H, Bohmer M, and Heierli C (1972) Serum

creatinine determination without protein precipitation.

Clinica Chimica Acta 37: 193–197.

Berkowitz GS, Wetmur JG, Birman-Deych E, Obel J,

Lapinski RH, Godbold JH, et al. (2004). In utero pesti-

cide exposure, maternal paraoxonase activity, and head

circumference. Environmental Health Perspectives

112(3): 388–391.

Brown SS, Kalow W, Pilz W, Whittaker M, and Woronick

CL (1981) The plasma cholinesterases; a new perspective.

Advances in Clinical Chemistry 22: 82–83.

Bulusu S, Chakravarty I (1986) Sub-acute administration of

organophosphorus pesticides and hepatic drug

metabolizing enzyme and malnourished rats. Bulletin of

Environmental Contamination and Toxicology 36: 73–80.

Butte NF, Garza C, Stuff JE, Smith EO, and Nichols BO

(1984) Effect of maternal diet and body composition

on lactational performance. American Journal of

Clinical Nutrition 39: 296–306.

Caglar C, Kolankaya D (2007) The effect of sub-acute and

sub-chronic exposure of rats to the glyphosate-based

herbicide Roundup. Environmental Toxicology and

Pharmacology 25: 57–62.

Collaborative Group on Hormonal Factors in Breast Cancer

(2002) Breast cancer and breastfeeding: collaborative

reanalysis of individual data from 47 epidemiological

studies in 30 countries, including 50302 women with

breast cancer and 96973 without the disease. Lancet

360: 187–195.

Conolly RB, Beck BD, and Goodman JI (1999) Stimulating

research to improve the scientific basis of risk assess-

ment. Toxicology Sciences 49: 1–4.

Cook JC, Kaplan AM, Davis LG, and O’Connor JC (1997)

Development of a Tier I screening battery for detecting

endocrine-active compounds (EACs). Regulatory

Toxicology Pharmacology 26: 60–68.

Davis DL, Ahmed AK (1998) Exposures from indoor

spraying of chlorpyrifos pose greater health risks to

children than currently estimated. Environmental Health

Perspectives 106(6): 299–301.

de Boer WB, Segal A, Frost FA, and Sterrett GF (2000)

Can CD34 discriminate between benign and malignant

hepatocytic lesions in fine-needle aspirates and thin core

biopsies? Cancer 90: 273–278.

De Kanter R, De Jager MH, Draaisma AL, Jurva, JU,

Olinga P, Meijer DK, et al. (2002) Drug-metabolizing

activity of human and rat liver, lung, kidney and intes-

tine slices. Xenobiotica 32: 349–362.

Draper HH, Hadley M (1990) Malondialdehyde determina-

tion as index of lipid peroxidation. Methods in Enzymol-

ogy 186: 421–431.

El Sakka S, Salem E, and Abdel R (2002) In vitro hepato-

toxicity of alachlor and its by-products. Journal of

Applied Toxicology 22: 31–35.

Ellman GL, Courtney KD, Andres KD, and Featherstone

RM (1961) A new and rapid calorimetric determination

of acetylcholinesterase activity. Biochemical Pharma-

cology 7: 88–95.

Eskenazi B, Harley K, Bradman A, Weltzien E, Jewell NP,

Barr DB, et al. (2004) Association of in utero organo-

phosphate pesticide exposure and fetal growth and

length of gestation in an agricultural population. Envi-

ronmental Health Perspectives 112(10): 1116–1124.

Fishbeck KL, Rasmussen KM (1987) Effect of repeated

cycles on maternal nutritional status, lactational

performance and litter growth in ad libitum fed and

chronically food. Journal of Nutrition 117: 1967–1975.

Gallenberg LA, Vodicnik MJ (1989) Transfer of persistent

chemicals in milk. Drug Metabolism Review 21:

277–317.

Gill TS, Pande J, and Tewari H (1990) Sublethal e€ects of an

organophosphorus insecticide on certain metabolite lev-

els in a freshwater fish, Puntius conchonius Hamilton.

Pesticide Biochemistry and Physiology 36: 290–299.

Gill TS, Pande J, Tewari H (1991) Individual and combined

toxicity of common pesticides to teleost Puntius concho-

nius Hamilton. Indian Journal of Experimental Biology

29: 145–148.

Goel A, Dani V, and Dhawan DK (2005) Protective effects

of zinc on lipid peroxidation, antioxidant enzymes and

hepatic histoarchitecture in chlorpyrifos-induced

toxicity. Chemico-Biological Interactions 156: 131–140.

Goel A, Dani V, and Dhawan DK (2007) Zinc mediates

normalization of hepatic drug metabolizing enzymes

in chlorpyrifos-induced toxicity. Toxicology Letter

169: 26–33.

Gornall AC, Bardawill CJ, and David MM (1949) Determi-

nation of serum protein by means of biuret reaction.

Journal of Biological Chemistry 177: 751–756.

Gurunathan S, Robson M, Freeman N, Buckley B, Roy A,

Meyer R, et al. (1998) Accumulation of chlorpyrifos on


residential surfaces and toys accessible to children.

Environmental Health Perspectives 106(1): 9–16.

Habig WH, Pabst MJ, and Jakoby WB (1974) Glutathione

S-transferases, the first enzymatic step in mercapturic

acid formation. Journal of Biological Chemistry 249:

7130–7139.

Halliwell B (1996) Antioxidants in human health and

disease. Annual Review Nutrition 16: 33–50.

Halliwell B, and Gutteridge JMC (1999) Free Radicals in

Biology and Medicine. 3rd ed. New York: Oxford

University Press.

Hayes D, Pulford D (1995) The glutathione S-transferase

supergene family: regulation of GST and the contribu-

tion of the isoenzymes to cancer chemoprotection and

drug resistance. Critical Reviews in Biochemistry and

Molecular Biology 30: 445–600.

Interagency Staff Group on Carcinogens (1986) Chemical

carcinogens: a review of the science and its associated prin-

ciples. Environmental Health Perspectives 67: 201–282.

Kamataki T, Neal RA (1976) Metabolism of diethyl

p-nitrophenyl phosphorothionate (parathion) by a recon-

stituted mixed-function oxidase enzyme system: studies

of the covalent binding of the sulfur atom. Molecular

Pharmacology 12: 933–944.

Kerem M, Bedirli N, Gurbuz N, Ekincl O, Bedirli A,

Akkaya T, et al. (2007) Effects of acute fenthion toxicity

on liver and kidney function and histology in rats.

Turkish Journal of Medical Sciences 37: 281–288.

Khan SM, Sobti RC, and Kataria L (2005) Pesticide-

induced alteration in mice hepato-oxidative status and

protective effects of black tea extract. Clinica Chimica

Acta 358: 131–138.

Kramer MS, Kakuma R (2002) Optimal duration of exclu-

sive breastfeeding. Cochrane Database of Systematic

Reviews; (7): CD003517.

Kuester RK, Waalkes MP, Goering PL, Fisher BL,

McCuskey RS, and Sipes IG (2002) Differential hepato-

toxicity induced by cadmium in Fischer 344 and

Sprague–Dawley rats. Toxicology Sciences 65: 151–159.

Mahboob M, Siddiqui MK (2001) Alterations in hepatic

detoxifying enzymes induced by neworganophosphorus

insecticides following subchronic exposure in rats.

Journal of Applied Toxicology 21: 501–505.

Mansour SA, Heikal TM, Mossa AH, and Refa A (2008)

Toxic effects of five insecticides and their mixture on

male albino rats. Journal of Egyptian Society of Toxicology.

39: 85–94.

Mansour SA, Mossa AH (2005) Comparative effects of

some insecticides as technical and formulated on male

albino rats. Journal of Egyptian Society of Toxicology

32: 41–54.

Mansour SA, Mossa AH (2009) Lipid peroxidation and

oxidative stress in rat erythrocytes induced by chlorpyr-

ifos and the protective effect of zinc. Pesticide Biochem-

istry and Physiology 93: 34–39.

Mansour SA, Mossa AH (2010a) Oxidative damage,

biochemical and histopathological alterations in rats

exposed to chlorpyrifos and the antioxidant role of zinc.

Pesticide Biochemistry and Physiology 96: 14–23.

Mansour SA, Mossa AH (2010b) Adverse effects of

lactational exposure to chlorpyrifos in suckling rats.

Human and Experimental Toxicology 92(2): 77–92.

Marty MS, Domoradzki JY, Hansen SC, Timchalk C,

Bartels MJ, and Mattsson JL (2007) The effect of route,

vehicle, and divided doses on the pharmacokinetics of

chlorpyrifos and its metabolite trichloropyridinol in

neonatal Sprague-dawley rats. Toxicology Science 100:

360–373.

Mattsson JL, Maurissen JP, Nolan RJ, and Brzak KA

(2000) Lack of differential sensitivity to cholinesterase

inhibition in fetuses and neonates compared to dams

treated perinatally with chlorpyrifos. Toxicology

Sciences 53: 438–446.

Maurissen JPJ, Hoberman AM, Garman RH, and Hanley

TR Jr (2000) Lack of selective developmental neuro-

toxicity in rat pups from dams treated by gavage with

chlorpyrifos. Toxicology Sciences 57: 250–263.

McNamara JP, Hillers JK (1986) Adaptations in lipid

metabolism of bovine adipose tissue in lactogenesis and

lactation. Journal of Lipid Research 27: 150–157.

Moss DW, and Henderson AR (1999) Enzymes. In:

Burtis CA, Ashwood ER (eds.) Tietz Textbook of Clinical

Chemistry. 2nd ed. Philadelphia: WB Saunders, 735–896.

Murray KF, Messner DJ, and Kowdley KV (2006) Mechan-

isms of hepatocyte detoxification. In: Johnson LR (ed.)

Physiology of the Gastrointestinal Tract. Burlington:

Elsevier Academic Press, 1483–1503.

National Research Council (NRC) (1996) Guide for the

Care and Use of Laboratory Animals. Washington,

DC: Academic Press.

Navarro CM, Montilla PM, Martin A, Jimenez J, and

Utrilla PM (1993) Free radicals scavenger and

antihepatotoxic activity of Rosmarinus. Plant Medicine

59: 312–314.

Nolan RJ, Rick DL, Freshour NL, and Saunders JH (1984)

Chlorpyrifos: pharmacokinetics in human volunteers.

Toxicology and Applied Pharmacology 73: 8–15.

Peraino C, Richards WL, and Stevens J (1983) Multi-stage

hepatocarcinogenesis. Environmental Health Perspec-

tives 56: 1–43.

Ranjbar A, Pasalar P, and Abdollahi M (2002) Inducation

of oxidative stress and acetylcholinesterase inhibition


in organophosphorous pesticide manufacturing workers.

Human and Experimental Toxicology 21: 179–182.

Reitman S, Frankel S (1957) A colorimetric method for the

determination of serum glutamic oxalacetic and

glutamic pyruvic transaminases. American Journal of

Clinical Pathology 28: 56–63.

Rosenblatt KA, Thomas DB (1993) WHO collaborative

study on neoplasia and steroid contraceptives. Interna-

tional Journal of Epidemiology 22: 192–197.

Salas JH, Gonzalez MM, Noa M, Perez NA, Dıaz G,

Gutierrez R, et al. (2003) Organophosphorus pesticides

in Mexico commercial pasteurized milk. Journal of

Agriculture and Food Chemistry 51: 4468–4471.

Sanghi R, Pillai MK, Jayalekshmi TR, and Nair A (2003)

Organochlorine and organophosphorus pesticides resi-

dues in breast milk from Bhopal, Madhya Pradesh,

Indian. Human and Experimental Toxicology 22: 73–76.

Sevanian A, Nordenbrand K, Kim E, Ernster L, and

Hochstein P (19900 Microsomal lipid peroxidation:

the role of NADPH-cytochrome P450 reductase and

cytochrome P450. Free Radical Biology and Medicine

8: 145–152.

Shakoori AR, Butt U, Riffat R, Aziz F (1994) Hematologi-

cal and biochemical effects of danitol administered for

two months on the blood and liver of rabbits. Zeitschrift

fuer Angewandte Zoologie 80: 165–180.

Szasz G (1969) Kinetic photometric methods for serum

gamma glutamyltranspeptidase. Clinical Chemistry 15:

124–136.

Tietz NM (1995) Clinical Guide to Laboratory Tests. Phi-

ladelphia, PA: WB Saunders Company, 384–385.

Tomlin CDS (2005) The e-Pesticide Manual. 13th ed.

CDROM version 3.1, The British Crop Protection

Councill: Farnham, UK.

Wang X, Zhai W (1988) Cellular and biochemical

in bronchoalveolar lavage fluids of rats exposed to

fenvalerate. Zhongguo Yaolixue YuDulixue Zoghi 2:

271–276.

Westgard JO, Poquette MA (1972) Determination of

serum albumin with the ‘‘SMA 12-60’’ by a bromcresol

green dye-binding method. Clinical Chemistry 18:

647–653.

Williams GM, Iatropoulos MJ (2002) Alteration of

liver cell function and proliferation: differentiation

between adaptation and toxicity. Toxicology Pathol-

ogy 30: 41–53.

Woolliams JA, Wiener G, Anderson PH, and McMurray CH

(1983) Variation in the activities of glutathione

peroxidase and superoxide dismutase and in the concen-

tration of copper in the blood various breed crosses of

sheep. Research in Veterinary Science 34: 69–77.

Yamano T, Morita S (1992) Hepatotoxicity of trichlorfon

and dichlorvos in isolated rat hepatocytes. Toxicology

76: 69–77.

Yano BL, Young JT, and Mattsson JL (2000) Lack of

carcinogenicity of chlorpyrifos insecticide in a high-dose,

2-year dietary toxicity study in Fischer 344 rats. Toxicology

Sciences 53: 135–144.


Adverse effects of exposure to low doses of chlorpyrifos in lactating rats

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