UNITED NATIONS ENVIRONMENT PROGRAMME
INTERNATIONAL LABOUR ORGANISATION
WORLD HEALTH ORGANIZATION
INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY
ENVIRONMENTAL HEALTH CRITERIA 198
Diazinon
This report contains the collective views of an international group of
experts and does not necessarily represent the decisions or the stated
policy of the United Nations Environment Programme, the International
Labour Organisation, or the World Health Organization.
Environmental Health Criteria 198
First draft prepared by Dr K. Barabás, Albert Szent-Gyorgyi University
Medical School, Szeged, Hungary
Published under the joint sponsorship of the United Nations
Environment Programme, the International Labour Organisation, and the
World Health Organization, and produced within the framework of the
Inter-Organization Programme for the Sound Management of Chemicals.
World Health Organization
Geneva, 1998
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of the biological action of chemicals.
WHO Library Cataloguing in Publication Data
Diazinon
(Environmental health criteria ; 198)
1.Diazinon - toxicity 2.Diazinon - adverse effects
3.Environmental exposure 4.Occupational exposure
I.International Programme on Chemical Safety II.Series
ISBN 92 4 157198 5 (NLM Classification: WA 240)
ISSN 0250-863X
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CONTENTS
ENVIRONMENTAL HEALTH CRITERIA FOR DIAZINON
PREAMBLE
ABBREVIATIONS
1. SUMMARY
1.1. Identity, physical and chemical properties, analytical
methods
1.2. Production, uses and sources of human and environmental
exposure
1.3. Environmental transport, distribution and transformation
1.4. Environmental levels and human exposure
1.5. Kinetics and metabolism
1.6. Effects on experimental animals and in vitro test systems
1.7. Effects on humans
1.8. Effects on other organisms in the laboratory and field
2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, ANALYTICAL METHODS
2.1. Identity
2.1.1. Primary constituent
2.1.2. Technical product
2.2. Physical and chemical properties
2.3. Analytical methods
3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE
3.1. Natural occurrence
3.2. Man-made sources
3.2.1. Production levels and processes
3.2.1.1 Manufacturing process
3.2.2. Uses
3.2.3. Formulations
4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION
4.1. Transport and distribution between media
4.1.1. Volatilization
4.1.2. Movement in soil
4.2. Degradation
4.2.1. Degradation in soil
4.2.2. Degradation in water
4.2.3. Bioconcentration
4.2.3.1 Fish and aquatic invertebrates
5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE
5.1. Environmental levels
5.1.1. Air
5.1.2. Water
5.1.3. Soil
5.1.4. Fruit, vegetables and food
5.1.5. Milk
5.1.6. Meat and fat
5.2. General population exposure
5.3. Occupational exposure
6. KINETICS AND METABOLISM
6.1. Absorption, distribution and excretion
6.1.1. Oral administration
6.1.1.1 Rats
6.1.1.2 Guinea-pigs
6.1.1.3 Dogs
6.1.1.4 Goats
6.1.1.5 Cow
6.1.1.6 Hens
6.1.2. Dermal application
6.1.2.1 Rats
6.1.2.2 Sheep
6.1.2.3 Humans
6.1.3. Other routes
6.1.3.1 Intraperitoneal administration
6.1.3.2 Subcutaneous administration
6.1.3.3 Intravenous administration
6.2. Metabolism
6.2.1. In vivo metabolic transformations
6.2.1.1 Mice
6.2.1.2 Rats
6.2.1.3 Dogs
6.2.1.4 Sheep
6.2.1.5 Goats
6.2.1.6 Hens
6.2.2. In vitro metabolic transformations
6.3. Metabolic aspects of diazinon toxicity
7. EFFECTS ON LABORATORY MAMMALS AND IN VITRO TEST SYSTEMS
7.1. Single exposure
7.1.1. Oral
7.1.2. Dermal
7.1.3. Inhalation
7.1.4. Intraperitoneal
7.2. Short-term exposure
7.2.1. Oral
7.2.1.1 Rats
7.2.1.2 Dogs
7.2.1.3 Pigs
7.2.2. Inhalation
7.2.3. Dermal
7.2.3.1 Rabbits
7.3. Long-term exposure
7.3.1. Rats
7.3.2. Dogs
7.3.3. Rhesus monkeys
7.4. Skin and eye irritation; sensitization
7.4.1. Primary skin irritation
7.4.2. Primary eye irritation
7.4.3. Skin sensitization
7.5. Reproduction, embryotoxicity and teratogenicity
7.5.1. Reproduction
7.5.1.1 Rat
7.5.1.2 Cattle
7.5.2. Embryotoxicity and teratogenicity
7.5.2.1 Mice
7.5.2.2 Rats
7.5.2.3 Hamsters
7.5.2.4 Rabbits
7.5.2.5 Chicken
7.6. Mutagenicity and related end-points
7.7. Carcinogenicity
7.7.1. Mice
7.7.2. Rats
7.8. Special studies
7.8.1. Neurotoxicity
7.8.2. Effects on enzymes and transmitters
7.8.3. Effects on the immune system
7.8.4. Effect on pancreas
7.9. Factors that modify toxicity; toxicity of metabolites
7.9.1. Metabolic enzymes
7.9.2. Antidotes
7.9.3. Potentiation
8. EFFECTS ON HUMANS
8.1. Exposure of the general population
8.1.1. Acute toxicity, poisoning incidents
8.1.1.1 Acute pancreatitis
8.1.1.2 Intermediate syndrome
8.1.1.3 Unusual case reports
8.1.2. Controlled human studies
8.2. Occupational exposure
8.2.1. Acute poisoning
8.2.2. Effect of short-term and long-term
exposure
9. EFFECTS ON OTHER ORGANISMS IN THE LABORATORY AND FIELD
9.1. Microorganisms
9.2. Aquatic invertebrates
9.3. Fish
9.4. Effects in mesocosms and the field
9.5. Terrestrial invertebrates
9.6. Birds
9.6.1. Field studies
10. EVALUATION OF HUMAN HEALTH RISK AND EFFECTS ON THE ENVIRONMENT
10.1. Evaluation of human health risk
10.2. Evaluation for effects on the environment
10.2.1. Aquatic organisms
10.2.1.1 Acute risk
10.2.1.2 Chronic risk
10.2.2. Terrestrial organisms
10.2.2.1 Birds
10.2.2.2 Mammals
10.2.2.3 Bees
10.2.2.4 Earthworms
11. CONCLUSIONS AND RECOMMENDATIONS FOR PROTECTION OF HUMAN HEALTH
AND THE ENVIRONMENT
11.1. Conclusions
11.2. Recommendations for protection of human health and the
environment
11.2.1. Recommendation on regulation of compound
11.2.1.1 Transport and storage
11.2.1.2 Handling
11.2.1.3 Disposal
11.2.1.4 Selection, training and medical
supervision of workers
11.2.1.5 Labelling
11.2.1.6 Residues in food
11.2.2. Prevention of poisoning in man and emergency aid
11.2.2.1 Manufacture and formulation
11.2.2.2 Mixers and applicators
11.2.2.3 Other associated workers
11.2.2.4 Other populations likely to be affected
11.2.3. Entry into treated areas
11.2.4. Emergency aid
11.2.5. Surveillance test
12. FURTHER RESEARCH
13. PREVIOUS EVALUATIONS BY INTERNATIONAL BODIES
REFERENCES
RÉSUMÉ ET ÉVALUATIONS
RESUMEN Y EVALUACIONES
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WHO TASK GROUP ON ENVIRONMENTAL HEALTH CRITERIA FOR DIAZINON
Members
Dr P.J. Abbott, Australia and New Zealand Food Authority
(ANZFA), Canberra, Australia
Dr K. Barabás, Department of Public Health, Albert Szent-Gyorgyi,
University Medical School, Szeged, Hungary
Dr A.L. Black, Woden, ACT, Australia
Professor J.F. Borzelleca, Pharmacology and Toxicology,
Richmond, Virginia, USA
Dr P.J. Campbell, Pesticides Safety Directorate, Ministry of
Agriculture, Fisheries and Food, Kings Pool, York,
United Kingdom
Professor L.G. Costa, Department of Environmental Health,
University of Washington, Seattle, USA
Dr S. Dobson, Institute of Terrestrial Ecology, Monks Wood,
Abbots Ripton, Huntingdon, Cambridgeshire, United Kingdom
Dr I. Dewhurst, Mammalian Toxicology Branch, Pesticides Safety
Directorate, Ministry of Agriculture, Fisheries and Food,
Kings Pool, York, United Kingdom
Dr V. Drevenkar, Institute for Medical Research and Occupational
Health, Zagreb, Croatia
Dr W. Erickson, Environmental Fate and Effects Division,
US Environmental Protection Agency, Washington, D.C., USA
Dr A. Finizio, Group of Ecotoxicology, Institute of Agricultural
Entomology, University of Milan, Milan, Italy
Mr K. Garvey, Office of Pesticide Programs (7501C),
US Environmental Protection Agency, Washington, D.C., USA
Dr A.B. Kocialski, Health Effects Division, Office of Pesticide
Programs, US Environmental Protection Agency,
Washington, D.C., USA
Dr A. Moretto, Institute of Occupational Medicine, University
of Padua, Padua, Italy
Professor O. Pelkonen, Department of Pharmacology and
Toxicology, University of Oulu, Oulu, Finland
Dr D. Ray, Medical Research Council Toxicology Unit, University
of Leicester, Leicester, United Kingdom
Dr J.H.M. Temmink, Department of Toxicology, Wageningen
Agricultural University, Wageningen, The Netherlands
Observers
Dr J.W. Adcock, AgrEvo UK Limited, Chesterford Park, Saffron,
Waldon, Essex, United Kingdom
Mr D. Arnold, Environmental Sciences, AgrEvo UK Ltd.,
Chesterford Park, Saffron Waldon, Essex, United Kingdom
Dr E. Bellet, CCII, Overland Park, Kansas, USA
Mr Jan Chart, AMVAC Chemical Corporation, Newport Beach,
California, USA
Dr H. Egli, Novartis Crop Protection AG, Basel, Switzerland
Dr P. Harvey, AgrEvo UK Ltd., Chesterford Park, Saffron Walden,
Essex, United Kingdom
Dr G. Krinke, Novartis Crop Protection AG, Basel, Switzerland
Dr A. McReath, DowElanco Limited, Letcombe Regis, Wantage,
Oxford, United Kingdom
Dr H. Scheffler, Novartis Crop Protection AG, Basel, Switzerland
Dr A.E. Smith, Novartis Crop Protection AG, Basel, Switzerland
Secretariat
Dr L. Harrison, Health and Safety Executive, Bootle, Merseyside,
United Kingdom
Dr J.L. Herrman, International Programme on Chemical Safety,
World Health Organization, Geneva, Switzerland
Dr P.G. Jenkins, International Programme on Chemical Safety,
World Health Organization, Geneva, Switzerland
Dr D. McGregor, Unit of Carcinogen Identification and Evaluation,
International Agency for Research on Cancer, Lyon, France
Dr R. Plestina, International Programme on Chemical Safety,
World Health Organization, Geneva, Switzerland
Dr E. Smith, International Programme on Chemical Safety,
World Health Organization, Geneva, Switzerland
Dr P. Toft, International Programme on Chemical Safety,
World Health Organization, Geneva, Switzerland
ENVIRONMENTAL HEALTH CRITERIA FOR DIAZINON
The Core Assessment Group (CAG) of the Joint Meeting on
Pesticides (JMP) met at the Institute for Environment and Health,
Leicester, United Kingdom, from 3 to 8 March 1997. Dr L.L. Smith
welcomed the participants on behalf of the Institute and
Dr R. Plestina on behalf of the three IPCS cooperating organizations
(UNEP/ILO/WHO). The CAG reviewed and revised the draft monograph and
made an evaluation of the risks for human health and the environment
from exposure to diazinon.
The first draft of the monograph was prepared by Dr K. Barabás,
Albert Szent-Gyorgyi University Medical School, Szeged, Hungary.
Extensive scientific comments were received following circulation of
the first draft to the IPCS contact points for Environmental Health
Criteria monographs and these comments were incorporated into the
second draft by the Secretariat.
Dr R. Plestina and Dr P.G. Jenkins, both members of the IPCS
Central Unit, were responsible for the overall scientific content and
technical editing, respectively.
The efforts of all who helped in the preparation and finalization
of the monograph are gratefully acknowledged.
ABBREVIATIONS
AChE acetylcholinesterase
ai active ingredient
ChE cholinesterase
CNS central nervous system
DETP diethylthiophosphate
DT degradation time
EDTA ethylenediaminetetraacetic acid
fc field capacity
GABA gamma-aminobutyric acid
ip intraperitoneal
MRL maximum residue limit
NAD nicotinamide adenine dinucleotide
NIOSH National Institute for Occupational Safety and Health (USA)
NOAEL no-observed-adverse-effect level
NOEC no-observed-effect concentration
NOEL no-observed-effect level
OSHA Occupational Safety and Health Administration (USA)
2-PAM pralidoxine (2-pyridine aldoxime methyl) chloride
PEC predicted environmental concentration
TEPP tetraethyl-pyrophosphate
TER toxicity-exposure ratio
TLV threshold limit value
1. SUMMARY
1.1 Identity, physical and chemical properties, analytical methods
The chemical name for diazinon is O, O-diethyl
O-2-isopropyl-6-methylpyrimidinyl-4-yl phosphorothioate. The pure
material forms a colourless liquid with a faint ester-like odour. The
technical active ingredient is a yellow/brown liquid with a slight
compound-specific odour. The boiling point is 83-84°C at 26.6 mPa and
the vapour pressure (volatility) is low (9.7 mPa at 20°C). The
solubility in water at room temperature is 60 mg/litre. Diazinon is
soluble in most organic solvents and has an octanol/water partition
coefficient (log Pow) of 3.40. It is stable in neutral media, but is
slowly hydrolysed in alkaline media and more rapidly in acid media. It
decomposes at temperatures above 120°C.
A large number of sampling and analytical methods have been
developed for the determination of diazinon and its metabolites in
different media. Sensitive methods, such as gas chromatography,
high-performance liquid chromatography, mass spectrometry and
immunoassay methods, are increasingly used.
1.2 Production, uses and sources of human and environmental exposure
Diazinon is a contact organophosphorus insecticide with a wide
range of insecticidal activity. It is effective against adult and
juvenile forms of flying insects, crawling insects, acarians and
spiders. It has been used from the early 1950s. Diazinon is mainly
formulated as wettable powders and emulsifiable concentrates. It is
also available in mixed formulations with other insecticides.
1.3 Environmental transport, distribution and transformation
Volatilization of diazinon from soil is of minor importance.
Diazinon has a tropospheric half-life of 1.5 h.
The movement of diazinon through soil is highly influenced by a
number of factors, particularly by organic matter and calcium
carbonate content. Diazinon is not expected to bind strongly to soil,
owing to its KOC value of 500, and is expected to show moderate
mobility in the soil.
Biological processes appear to be the main factor in the
degradation of diazinon in soil. At 20°C and a soil moisture content
of 60% of field capacity (f.c.) in a silt loam soil, the DT50 was
5 days. Under sterile conditions at 20°C and 60% f.c., the DT50 was
118 days, suggesting that biological activity is mainly responsible
for degradation in soil.
In natural water diazinon has a half-life of the order of 5-15
days. Both chemical and biological processes seem to play a role in
the degradation of diazinon, leading to mineralization within a few
weeks.
Uptake of diazinon by aquatic organisms is rapid. Low
bioconcentration factors have been reported for aquatic organisms,
ranging from 3 for shrimp to 152 for gudgeon, consistent with rapid
metabolism and loss. Depuration half-lives for fish have been reported
to be up to 30 h (muscle).
1.4 Environmental levels and human exposure
Environmental levels of diazinon are generally low. The routes of
exposure for the general population are inhalational and dietary.
Exposure through water is negligible. Occupational exposure is
primarily dermal.
Diazinon uses fall into two major categories: as a pesticide in
agriculture and as a drug in veterinary medicine. Thus, the major
source of diazinon residues in edible crops are from its use as an
agricultural pesticide, while those in meat, offal and other animal
products arise from its use as a veterinary drug containing active
ingredient.
Diazinon residues in vegetables, fruits and animal products are
very low. The results of total-diet studies suggest that diazinon
rapidly breaks down in both plant and animal products. Diazinon has
not been detected in drinking-water samples and its concentrations in
surface water are at the level of ng/litre.
1.5 Kinetics and metabolism
Diazinon may be absorbed from the gastrointestinal tract,
through the intact skin and following inhalation. Transdermal
absorption in humans is low. Diazinon is oxidized by the microsomal
enzymes to cholinesterase-inhibiting metabolites such as diazoxon,
hydroxydiazoxon, and hydroxydiazinon. Only minimal quantities of
metabolites are detectable in milk and eggs. Diazinon and its
metabolites do not accumulate in body tissue; 59-95% of an oral dose
of diazinon is excreted within 24 h and 95-98% is excreted within
7 days, mainly in urine.
The main metabolic pathways of degradation of diazinon are:
a) Cleavage of the ester bond leading to the hydroxypyrimidine
derivatives.
b) Transformation of P-S moiety to the P-O derivate.
c) Oxidation of isopropyl substituent leading to the corresponding
tertiary and primary alcohol derivatives.
d) Oxidation of the methyl substituent leading to the corresponding
alcohol.
e) Glutathione-mediated cleavage of the ester bond leading to a
glutathione conjugate.
The cleavage of the phosphorus ester bond, leading directly, or
via diazoxon, to the pyrimidyl metabolite plays the major role in the
metabolism of diazinon. Metabolites maintaining the phosphorus ester
bond are of transient nature and have been observed only in small
quantities. Yields and rates of production of metabolites vary greatly
between species. The production of diazoxon is not generally
correlated with susceptibility to diazinon poisoning, although it is
lowest in the least susceptible species, the sheep. The extrahepatic
metabolism of diazinon, especially the hydrolysis of diazoxon in
plasma, is more important toxicologically than the metabolism in
the liver, although the liver is probably the most important site
of metabolism in avian species. The metabolites formed, i.e.
diethylphosphoric acid, diethylthiophosphoric acid and the derivates
of the pyrimidinyl ring, are eliminated mainly via the kidneys.
1.6 Effects on experimental animals and in vitro test systems
Improvements in the manufacture of diazinon since 1979 have
significantly reduced the content of highly toxic impurities, e.g.,
tetraethyl-pyrophosphate (TEPP). As a result of these progressive
improvements, the acute oral LD50 of technical grade diazinon has
increased (e.g., from 250 mg/kg to 1250 mg/kg in the rat).
The acute oral, dermal and inhalational toxicity is low.
Short-term and long-term studies in mice, rats, rabbits, dogs and
monkeys have shown that the only effect of concern is dose-related
inhibition of acetyl cholinesterase activity.
Diazinon is slightly irritant to rabbit skin but not to the eye.
Diazinon is not a dermal sensitizer. Reproductive and developmental
studies have revealed no evidence of embryotoxic or teratogenic
potential. There was no effect on reproductive performance at dose
levels that were not toxic to the parent animals. Mutagenicity studies
with various end-points in vivo and in vitro gave no evidence of a
mutagenic potential. There is no evidence of carcinogenicity in rats
or mice. Diazinon does not cause delayed neuropathy in hens. In the
dog and guinea-pig, diazinon has been reported to cause acute
pancreatitis; this is considered to be a species-specific effect.
1.7 Effects on humans
Several cases of accidental or suicidal poisoning by diazinon
have been reported, some of which were fatal. In some of these the
cholinergic syndrome may have been more severe than expected because
of the presence of highly toxic impurities such as TEPP. In certain
cases, acute reversible pancreatitis was associated with a severe
cholinergic syndrome. This occurs also after poisoning with other
cholinesterase inhibitors. In a number of cases, the intermediate
syndrome was also observed. No cases of delayed neuropathy have ever
been reported, as expected from animal data. Reported cases of
poisoning after occupational exposure have always been associated with
the presence of impurities such as TEPP, monothio-TEPP or sulfo-TEPP
in the formulation. These impurities are unlikely to be found in
currently available formulations.
1.8 Effects on other organisms in the laboratory and field
Effects of diazinon on unicellular algae are variable; both
inhibition and stimulation of growth have been reported for different
species at concentrations between 0.01 and 5 mg/litre. Generally,
growth rates are reduced at concentration above 10 mg/litre, although
in certain cases population size can remain unaltered at 100 mg/litre.
Fewer and variable data make effects on other microorganisms difficult
to assess.
Acute LC50 values for aquatic invertebrates range from
0.2 µg/litre for Gammarus fasciatus to 4.0 µg/litre for the shrimp
Hyallela azteca in 96-h tests. Molluscs are substantially less
sensitive according to a single test on the snail Gillia
attilis. Sublethal effects on behaviour have been reported at
concentrations between 0.1 and 0.01 mg/litre.
Acute LC50 values for fish range from 0.09 mg/litre for rainbow
trout (Oncorhynchus mykiss) to 3.1 mg/litre for the catfish
(Channa punctatus). Growth of early life stages of fish was
inhibited at concentrations between 0.01 and 0.2 mg/litre. Brain
acetylcholinesterase activity is suppressed following acute exposure
to diazinon.
The LC50 for the earthworm Eisenia foetida in soil is
130 mg/kg soil.
The acute oral toxicity (LD50) in birds ranges from 1.1 mg/kg
body weight for Japanese quail to 85 mg/kg body weight for cowbirds.
Dietary LC50 values range from 32 mg/kg diet for mallard to 900 mg/kg
diet for Japanese quail (repellency was noted at these high dietary
concentrations). The no-observed-effect concentration in diet for
reproductive effects on birds in laboratory studies was 20 mg/kg
diet for mallard and 40 mg/kg diet for bobwhite quail. Brain
acetylcholinesterase activity is inhibited following ingestion.
Diazinon may also be taken in via the dermal route. There have been
reports of substantial field kills of water fowl following application
of diazinon to turf. Field studies applying liquid formulations to
turf at 4.8 kg ai/ha resulted in no mortality or reproductive effects
on song birds. Application of granules caused a small reduction in
song bird population size compared to that of controls. Ingestion of
small numbers of granules can be fatal for small birds, as
demonstrated in laboratory studies.
2. IDENTITY, PHYSICAL AND CHEMICAL PROPERTIES, ANALYTICAL METHODS
2.1 Identity
2.1.1 Primary constituent
Common name: diazinon
Chemical structure:
Chemical formula: C12H21N2O3PS
Relative molecular mass: 304.35
IUPAC Chemical names: O,O-diethyl O-2-isopropyl-6-methyl-
pyrimidin-4-yl phosphorothioate
CAS chemical name: O,O-diethyl O-[6-methyl-2-
(1-methylethyl)-4-pirimidinyl]
phosphorothioate
CAS registry number: 333-41-5
RTECS number: TF3325000
Official number: OMS 469; ENT 19 507
Synonyms: dimpylate, diazide, G.24480, Basudin,
Kayazinon, Necidol/Nucidol
2.1.2 Technical product
Trade names: Diazinon (Alpha, Darlingtons Mushroom
Laboratories, Murphy Chemicals and
Rentokil); Basudin (Ciba-Geigy);
Crompest (Cromessel Co. Ltd); Dethlac
(Gerhardt Pharmaceuticals); Isectalac
(Sorex Ltd); Murphy Root Guard (Fisons);
Rentokil Flytrol and Knox out 2FM
(Rentokil); Secto AntSpray and Root
Powder (Secto Ltd); Dazzel, Diagran,
Dianon (Nippon Kayaku); Diazotol
Gardentox, Nipsan (Nippon Kayaku);
Dyzol, Dizion (Nippon Kayaku);
Spectracide (Ciba-Geigy)
2.2 Physical and chemical properties
Diazinon is a clear colourless liquid (technical 95% yellow oil)
with a faint ester-like odour.
Boiling point: 83-84°C at 26.6 mPa; 125°C at 133 mPa
Vapour pressure: 9.7 mPa at 20°C
Density: 1.11 g/cm3 at 20°C
Refractive index: 1.4978-1.4981
Specific gravity: 1.116-1.118 at 20°C
Stability: susceptible to oxidation above 100°C;
stable in neutral media, but slowly
hydrolysed in alkaline media, and more
rapidly in acidic media
Decomposes: above 120°C
Corrosiveness: non-corrosive
Solubility: 60 mg/litre in water at 20°C;
completely miscible with common organic
solvents, e.g., ethers, acetone,
alcohols, benzene, toluene, cyclohexane,
hexane, dichloromethane, petroleum oils
2.3 Analytical methods
Formulated diazinon products are cleaned up by column
chromatography to remove the basic impurities and analysed by
titration with perchloric acid in acetic acid. They are also analysed
by gas-liquid chromatography (Eberle et al., 1974; Allender & Britt,
1994).
Residues in soil, water, air, plants, foods, and animal and human
tissues can be determined using gas chromatography using detectors
selective for phosphorus-containing compounds, and by other
chromatographic techniques. Table 1 outlines various methods for
determination of diazinon in different media.
Farran et al. (1988a) described a method for the determination of
organophosphorus insecticides and their hydrolysis products. The
method involves the analysis of compounds by liquid chromatography in
combination with UV and thermospray-mass spectrometric detection.
An automated identification method has been developed for water-
borne toxicants, including diazinon, using an ion chromatography/
high-performance liquid chromatography system (Fort et al., 1995).
A compendium of analytical methods for organophosphorus compounds
has been issued (NIOSH, 1994).
Table 1. Analytical methods for diazinon
Medium Analytical method References
Air adsorption on XAD-2 resin, gas NIOSH (1994)
chromatography with flame
photometric detector
Soil gas chromatography Singmaster & Acin-
Diaz (1991)
Water extraction with XAD-2 resin, gas Le Bel et al. (1979)
chromatography with nitrogen-
phosphorus detector, gas
chromatography/mass spectrometry
continuous-flow extraction coupled Farran et al. (1988b)
on-line with high-performance liquid
chromatography
liquid-solid extraction, gas Johnson et al. (1991)
chromatography/mass spectrometry
on-line solid-phase extraction, Lacorte & Barcelo
liquid chromatography/thermal spray - (1995)
mass spectrometry
on-line solid-phase extraction, Lacorte & Barcelo
liquid chromatography/atmospheric (1996)
pressure chemical ionization mass
spectrometry
maleic anhydride immunoassay Winnett (1992)
Oil solution gas chromatography Koibuchi et al.
(1975)
Fruit and solvent extraction, gas Ferreira & Silva
vegetables chromatography with thermionic Fernandes (1980)
detector
Apples solvent extraction, gas Asensio et al. (1991)
chromatography with thermionic
detector
Table 1. (con't)
Medium Analytical method References
Oranges matrix solid-phase dispersion Torres et al. (1996)
extraction, gas chromatography with
electron capture detector
Rice solvent extraction, gas Adachi et al. (1984)
chromatography with flame ionization
detector
Spinach preparative thin-layer Gilmore & Cortes
chromatography, autoradiography, (1996)
liquid scintillation counting
solvent extraction, gas Cairns et al. (1985)
chromatography with electrolytic
conductivity detector, gas
chromatography/chemical ionization
mass spectrometry
Milk gas chromatography Toyoda et al. (1990)
Human tissue solvent extraction, thin-layer Kirkbride (1987)
chromatography, gas chromatography
with nitrogen-phosphorus detector
Blood plasma gas chromatography Machin et al. (1975)
solvent extraction, gas Wu et al. (1994)
chromatography with electron capture
detector
Metabolites in urine
DEP, DEPT extraction by anion exchange resin, Lores & Bradway
gas chromatography with flame (1977)
photometric detector Weisskcp & Seiber
(1989)
GW7 550, solvent extraction, gas Lawrence & Iverson
GS 31 144 chromatography with electrolytic (1975)
conductivity detector
3. SOURCES OF HUMAN AND ENVIRONMENTAL EXPOSURE
3.1 Natural occurrence
Diazinon does not occur as a natural product.
3.2 Man-made sources
3.2.1 Production levels and processes
3.2.1.1 Manufacturing process
Diazinon is the common name for O, O-diethyl
O-2-isopropyl-6-methylpyrimidin-4-yl phosphorothioate (IUPAC name),
an organophosphate insecticide. Its insecticidal properties were first
described by Gasser (1953) and it was introduced in 1952, by
J. R. Geigy S.A. under the code number G 24480, trade names Basudin,
Diazitol, Neocidol and Nucidol, and the protection of BP 713278; USP
2754243. Meanwhile, improvements in the manufacturing process and the
stabilization of the technical grade diazinon by epoxidized soybean
oil have significantly reduced the content and formation of toxic
by-products and breakdown products and have reduced the acute toxicity
of diazinon products.
3.2.2 Uses
Diazinon is a contact organophosphorus insecticide with a wide
range of insecticidal activity, having long persistence and relatively
low mammalian toxicity. Diazinon is effective against adult and
juvenile forms of insects, but also against acarina. The spectrum of
activity includes the following arthropod groups:
* flying insects: flies and fly maggots, mosquitoes
* crawling insects: cockroaches, bedbugs, lice and ants
* acarina: dog ticks
* arachnideae: spiders
The main applications are rice, fruit, vineyards, sugar-cane,
corn, tobacco, potatoes, horticultural crops, animal dips and sprays.
Diazinon is also used by trained pest control operators in
households and outbuildings to control cockroaches, ants, silverfish,
spiders, carpet beetles and scorpions and in insecticidal collars on
domestic pets.
3.2.3 Formulations
The most important diazinon formulations are: ULV concentrates,
wettable powders 400 g/kg; emulsifiable concentrates 600, 400 and
250 g/litre; dust 20-40 g/kg; granules 30-140 g/kg; aerosols
200 g/litre.
Some typical formulations for agricultural and horticultural use
include: Basudin 5 (50 g a.i./kg); Basudin 10 (100 g a.i./kg) GR;
Basudin 40WP (400 g a.i./kg); Basudin 50SD (500 g a.i./kg); Basudin
60EC (EC 600 g a.i./litre); Diazitol Liquid; Basudin Ulvair 500;
Basudin 20 Mushroom Aerosol, KN; Knox-out (Pennwalt), flowable
microcapsules (230 g a.i./litre); Neocidol 60, Nucidol 60,
EC (600 g a.i./litre) for veterinary use.
4. ENVIRONMENTAL TRANSPORT, DISTRIBUTION AND TRANSFORMATION
4.1 Transport and distribution between media
4.1.1 Volatilization
It has been shown that diazinon is lost from soil through
volatilization (Harris & Mazurek, 1966), but the rate of loss is
unknown. Results of earlier studies with 14C-labelled insecticide and
the use of capped containers for holding treated soils indicated that
volatility was of minor importance. Under field conditions,
co-distillation, high temperatures and exposed surface areas probably
contribute to a greater loss of the insecticide through
volatilization.
Rate estimations according to the Atkinson incremental method
indicate that diazinon is rapidly degraded by hydroxyl radicals in the
atmosphere. The tropospheric half-life of diazinon lies between 1.3
and 1.5 h (Stamm, 1994).
4.1.2 Movement in soil
A study concerning degradation rate and mobility of diazinon in a
thatch layer of turf grass and in the underlying soil (2.5 cm) was
performed by Sears & Chapman (1979). Immediately following the
pesticide application, 2200 litres of water were applied to the total
treated area of 80 m2. Fourteen days after the application, less than
2% of the compound remained in the grass-thatch layer, and less than
1% in the root zone and in the underlying soil. The authors concluded
that the compound readily disappeared by degradation and/or
volatilization. However, it must be considered that only the top
2.5-cm layer was analysed.
The movement of diazinon and other organophosphorus compounds in
the soil was evaluated by means of soil thin-layer chromatography
(Sharma et al., 1986). The experiment was performed with two types of
soil (silt loam and sandy loam) showing different percentages of
organic matter (1.05 and 0.35%, respectively). The authors found a
generally poorer movement of diazinon in the silt loam soil, probably
due to the higher organic matter content and higher cation exchange
capacity. When natural soils were used as adsorbent and distilled
water as eluent, diazinon showed relatively high mobility. In this
study, the effects of pH and the presence of leachates of alkaline and
saline salts were also evaluated. Diazinon showed a slight decrease of
mobility in both soils at pH 4, whereas at pH 10 there was increase
mobility in the silt loam and slight decrease in mobility in the sandy
loam. The effects of leachate salt were not significant, with the
exception of calcium sulfate, which decreased mobility in the silt
loam soil.
The adsorption and mobility of diazinon in 25 Spanish soils and
the influence of soil properties on both processes were studied
(Arienzo et al., 1994). Adsorption constants of diazinon in the soils
were measured using soil thin-layer chromatography and soil column
leaching. The experiments were conducted with 14C-labelled diazinon.
Adsorption of diazinon was found to follow the Freundlich adsorption
equation. The Freundlich adsorption constant (K) ranged from 0.70 to
25.73. Adsorption was highly significantly correlated (p <0.001) with
the content of organic matter (OM). The median KOM value was 290
corresponding to a KOC value of 500. There was also a significant
correlation (p <0.01) of K and the distribution coefficient Kd with
the silt-plus-clay content in soils with low organic material content
(<2%). On the basis of the soil thin-layer chromatography
experiments, diazinon was found to be slightly mobile in 80% and
immobile in 20% of the soils studied. In the soil column experiment,
the pesticide was quite mobile under saturated flow in soils of light
texture containing little organic matter. Under non-saturated flow
conditions, which are more similar to natural conditions, diazinon
should not be easily leached from the studied soils to groundwater.
4.2 Degradation
4.2.1 Degradation in soil
Seyfried (1994) studied the degradation of diazinon in an
agricultural soil (silt loam, USDA) under various experimental
conditions. At 20°C and a soil moisture of 60% of the field capacity,
the DT50 was 5 days and DT90 22 days. The main metabolite,
2-isopropyl-4-methyl-6-hydroxy pyrimidine, occurred transiently and
degraded with a DT50 of 20 days. Mineralization accounted for 86% of
the applied diazinon within the experimental period of 120 days.
Whereas the application rate did not influence the degradation rate,
there was a dependence on temperature (DT50 of 12 days at 10°C) and
soil moisture (DT50 of 8 days at 30% field capacity). Under sterile
conditions, the DT50 was increased to 118 days at 20°C and 60% field
capacity. This suggests that the main route of soil degradation is
microbial.
Getzin (1968) studied persistence of diazinon in soils and
measured loss in autoclaved and non-autoclaved soil at several
temperatures, moisture contents and pH levels under controlled
laboratory conditions. Microorganisms and non-biological factors
affected the persistence of diazinon in Sultan silt loam. Diazinon was
primarily degraded through non-biological pathways. Although diazinon
was not metabolized to any great extent by microorganisms in Sultan
silt loam, it is known that soil microflora are capable of degrading
the insecticide. Gunner et al. (1966) isolated a bacterium from soil
that utilized diazinon as a source of sulfur, phosphorus, carbon and
nitrogen, but the importance of this microorganism as a contributor to
the metabolism of the insecticide in soil was not determined.
Miles et al. (1978) demonstrated that diazinon can accumulate and
persist in organic soils for more than a year. It was also shown that
diazinon can move from its soil-bound form into the aqueous
environment either via leaching or by direct soil erosion (Miles &
Harris, 1978a). Morganian & Wall (1972) demonstrated that diazinon
treatment of a marine salt marsh led to a build-up of diazinon in salt
marsh sod and mud.
At pH 6.8, the time required for 50% loss of diazinon is 6 weeks
in autoclaved soil and 18 weeks in buffered water. Mortland & Raman
(1967) demonstrated the catalytic hydrolysis of diazinon in CuCl2
solutions and Cu-montmorillonite suspensions. Catalytic reactions of
this nature may occur in soil, but attempts to demonstrate this
phenomenon in Sultan silt loam have so far failed. Moisture variations
from 50 to 100% of the moisture equivalent did not appreciably alter
the degradation rates of diazinon. Variations in soil temperature
between 10 and 30°C resulted in a 4- to 10-fold difference in the time
required for 50% loss of the insecticides in soil. The non-biological
degradation of diazinon increased with increased acidity.
Schoen & Winterlin (1987) have studied the factors affecting the
rate of diazinon degradation in soil. These are pH, soil type, organic
amendments, soil moisture and pesticide concentration. Of the soil
factors investigated, the conditions for diazinon degradation in
pesticide mixtures were optimum when the pesticides were present at
low concentrations in moist soil, amended with peat and acidified to
pH 4. Degradation was least at high pesticide concentration in neutral
or alkaline mineral soil.
Utilization of diazinon by an Arthrobacter species and a
Streptomyces species has been shown to alter the microbial
population by stimulating a selective enrichment of these species.
The Arthrobacter species previously reported to attack the side
chain of the molecule was unable to metabolize completely the ring
portion of the molecule. Similar results demonstrated that the
Streptomyces species, too, could not by itself convert pyrimidinyl
carbon to carbon dioxide. When, however, both the Arthrobacter and
Streptomyces organisms were incubated together, 15-20% of the 14C
appeared as labelled BaCO3 after 18 h, suggesting a synergistic
relationship between these two organisms in attacking the pyrimidinyl
portion of diazinon (Gunner & Zuckerman, 1968).
Barik & Munnecke (1982) demonstrated that a bacterial enzyme can
hydrolyse diazinon in soil. In their research, an enzyme was obtained
from a Pseudomonas sp. that could hydrolyse diazinon and several
other methoxy- or ethoxy-substituted organophosphates. In this
experiment, diazinon, either in 25% EC formulations or as a technical
grade chemical, was enzymatically hydrolysed in an agricultural sandy
soil when present at concentrations up to 1%. The degradation rate was
approximately proportional to enzyme concentration up to 12 units per
20 g soil. This indicates that the initial rate of diazinon
degradation is directly dependent on enzyme activities, and not on
chemical or physical parameters of the soil-pesticide interactions.
Although the enzyme was examined only in one soil, it is expected that
it could also operate on cement or asphalt type surfaces, as well as
on synthetic polymers such as carpet.
Al-Attar & Knowles (1982) studied the uptake, metabolism and
elimination of diazinon in Panagrellus redivivus, a free-living soil
nematode, and Bursaphelenchus xylophilus, a plant parasitic
nematode. Nematodes were exposed to a solution of diazinon labelled
with radiocarbon. Both nematode species metabolized diazinon, although
P. redivivus was more active. Metabolites from B. xylophilus
included O, O-diethyl O-(2-isopropyl-4-methyl-6-pyrimidinyl)
phosphate or diazoxon and pyrimidinol. Radioactivity accumulated to a
greater extent in B. xylophilus than in P. redivivus. Elimination
of radiocarbon was more rapid with P. redivivus than with
B. xylophilus, and this resulted in the presence of high levels
of the polar pyrimidinol metabolite in the incubation medium of
P. redivivus.
4.2.2 Degradation in water
Keller (1983) investigated the degradation of diazinon in samples
of pond and river water, each containing 1% of sediment. Diazinon was
degraded with a DT50 of 7 to 10 days in the pond system and 8 to 15
days in the river water. Mineralization accounted for >60% of the
applied material within 7 weeks in both systems.
In a mesocosm study conducted with 17 treated and 4 untreated
ponds (0.05 hectare each), diazinon degraded rapidly. The
disappearance half-lives averaged 5.2 to 12.2 days (Giddings, 1992).
Kanazawa (1975) found diazinon to be fairly persistent in tap
water in a glass aquarium, degrading to 27% in 30 days.
Ferrando et al. (1992) studied the persistence of diazinon in
natural water from Albufera Lake and in experimental water from their
laboratory. Degradation was faster in lake water, the half-lives being
70 and 79 h for lake and laboratory water, respectively. The
degradation process in both media was comparable until 96 h. The
authors found 43.5 and 49.4% of the applied diazinon in natural and
experimental water, respectively, at 96 h.
4.2.3 Bioconcentration
4.2.3.1 Fish and aquatic invertebrates
The bioconcentration factors (BCF) of diazinon over a 7-day
period were as follows: topmouth gudgeon 152; carp 65; guppy 18;
crayfish 4.9; red snail 17; pond snail 5.9 (Kanazawa, 1978).
Seguchi & Asaka (1981) reported the intake and excretion of
diazinon and its metabolites in freshwater fish, and the relationship
between the BCF of diazinon and fat content of fish. During exposure
to continuous-flow water containing 0.02 mg diazinon/litre the
concentration of diazinon in fish rapidly increased, reaching a
maximum after 3 days. Thereafter, the diazinon concentration slightly
decreased and remained at equilibrium. The BCFs for carp, rainbow
trout, leech and shrimp at equilibrium were 120, 63, 26 and 3,
respectively. As for the metabolites, pyrimidine analogue was found in
all fish species, but diazinon and related compounds were found only
in carp and rainbow trout. The concentration of the metabolites
reached a maximum after 3-7 days exposure to diazinon. Diazinon was
metabolized to diazoxon in the channel catfish liver microsomal enzyme
system, but it was not found in any other fish species. When the fish
were transferred to clean water, diazinon and its metabolites were
rapidly lost from the fish. Seven days after being transferred to
clean water, the diazinon concentration decreased to 0.3-8.0% of the
equilibrium concentration, and the metabolites decreased below the
detection limit.
Similar results have been observed for topmouth gudgeon by
Kanazawa (1975, 1978). A linear relationship was observed between the
bioconcentration ratio and fat content in fish. Seguchi & Asaka (1981)
identified six metabolites of diazinon, and Fujii & Asaka (1982)
identified another three: hydroxydiazinon, hydroxymethyl diazinon and
isopropenyl diazoxon.
The toxicity, accumulation and elimination of diazinon were
investigated in the European eel (Anguilla anguilla). Fish exposed
to sublethal concentration (0.042 mg/litre) accumulated diazinon in
the liver and muscle tissues. The BCFs for diazinon were 1859 in liver
and 775 in muscle over the 96-h exposure period. When removed from
diazinon-containing water, the contaminated fish rapidly eliminated
diazinon. The excretion rate constants were 0.108 per h for liver and
0.016 per h for muscle. Diazinon half-lives were 16.6 and 33.2 h for
liver and muscle, respectively (Sancho et al., 1992).
The freshwater fish Motsugo (Pseudorasbora parva) was reared in
an aquarium tank containing about 1 mg diazinon/litre for 30 days. The
persistence of the insecticide in water and the uptake and excretion
of the insecticide by fish were monitored. Diazinon degraded by 72% in
30 days. The concentration of diazinon in fish reached a maximum level
of 211 mg/kg after 3 days. Afterwards, the concentration of the
insecticide decreased gradually due to metabolism and excretion
(Kanazawa, 1975).
Bioconcentration and excretion of diazinon were studied in the
carp ( Cyprinus carpio L.). The average BCF values for diazinon were
20.9 in muscle, 60.0 in liver, 111.1 in kidney and 32.2 in gall
bladder over a 168-h exposure period. The excretion rate constants of
diazinon (ng/g per h) were 0.002-0.024 for muscle, 0.001-0.020 for
liver, 0.0004-0.004 for kidney and 0.002-0.023 for gall bladder,
respectively (Tsuda et al., 1990).
5. ENVIRONMENTAL LEVELS AND HUMAN EXPOSURE
5.1 Environmental levels
5.1.1 Air
The amount of insecticide present in the air of commercial pest
control buildings, service vehicles and food preparation-serving areas
following routine commercial insecticide application has been
measured. Diazinon was measured in the ambient air of storage and
office rooms in six North Carolina (USA) firms in a 4-h period. In the
storage rooms the mean value was 284 (85-837) ng/m3 air and in the
offices 163 (31-572) ng/m3 air. Diazinon was also detected in the
ambient air of vehicles used in commercial pest control activities.
Mean diazinon concentrations (ng/m3 air) in 2-h of sampling from six
vehicles were 88 (7-239) in sedans and 171 (11-543) in vans, the mean
value being 130 (7-543). The highest level of diazinon detected in the
ambient air of offices of pest control building was far below the
allowable limits (TLV : 100 µg/m3) (Wright & Leidy, 1980).
Wright et al. (1982) studied the amount of diazinon in cabs of
stationary pick-up trucks used by the pest control service. Additional
air samples, taken while the same pick-up truck was moving, provided
data for comparison of insecticide levels in individual pick-up trucks
when moving and stationary. Diazinon was present in significantly
greater concentrations than chlorpyrifos. This may be attributable to
the facts that the service technicians kept diazinon in sprayers
during the sampling periods and that they used it in servicing
accounts during the sampling day. It could therefore have contaminated
their clothing and skin and passed into the air when they were in the
pick-ups. The maximum diazinon detected was 5.15 µg/m3 for a 2-h
period or 20.6 µg/m3 for an 8-h period, which is about 1/5 of the
allowable limit. However, the amount of airborne diazinon to which a
technician was actually exposed during a working day was even less
than 20.6 µg/m3, since the maximum time any technician spent in a
pick-up was 3.8 h.
Wachs et al. (1983) reported the concentration of diazinon in
the air of a retail garden store that sold the insecticide. The
concentration found in the air based on the 14-h period pumping
through the polyurethane filters was 3.4 µg/m3. All of the diazinon
was found in the polyurethane plug closest to the air inlet. Diazinon
was not found in the second plug or unused plugs which were similarly
Soxhlet-extracted and analysed. It was concluded that the
concentration of diazinon in air depended on a number of factors,
including the type of formulation, air temperature, type and condition
of containers, prior spills and types of floor covering. The
concentration of diazinon vapour found in this study would not appear
to constitute a hazard to store personnel or customers.
Airborne concentrations of diazinon were measured in rooms for
21 days after crack and crevice application. Residue levels were
greatest in treated rooms (38 µg/m3) followed by adjacent (1 µg/m3)
and upper and lower floor rooms (about 0.4 µg/m3). Low levels of
diazinon were detected in all rooms 21 days after application. Small
amounts of diazinon (corrected to an 8-min application period) were
detected on respirator pads (2.6 µg) and waist pads (2.3 µg) worn by
the applicator (Leidy et al., 1982).
Airborne and surface concentrations of diazinon were measured at
intervals up to 10 days after broadcast spray application onto the
floors of seven offices. Diazinon concentrations peaked 4 h after
application at 163 and 27 µg/m3 of air sampled, respectively.
Airborne concentrations of diazinon indicated that building occupants
should not enter unventilated rooms for at least 2 days after
spraying. Residues on aluminium plates and furniture were examined at
intervals of up to 48 h after spraying, and in many cases the surface
concentrations were higher at 24 or 48 h after spraying than at
one hour. The peak residue concentration of diazinon was 38 ng/cm2 of
surface area sampled at 48 h (Currie et al., 1990).
5.1.2 Water
Insecticide residues on suspended and bottom sediments of streams
of Ontario, Canada, have been studied in a tobacco-growing, vegetable
muck area. Bed load samples contained three to six times higher
concentrations of insecticides than bottom material (Miles, 1976).
From 1985 to 1987, a monitoring survey was conducted to determine
the levels of selected pesticides in farm ditches located in the lower
mainland of British Columbia, Canada. Diazinon was not detected in
ditch water (detection limit = 1 µg/litre). In ditch sediments,
diazinon was sporadically found at concentrations up to 4 µg/kg
(detection limit = 1 µg/kg) (Wan, 1989).
During the first half of 1984, diazinon was not detected in raw
or treated water samples from the Lakeview and Lorne Park Water
Treatment Plants in Toronto, Ontario (detection limit = 10 ng/litre)
(MacLaren Plansearch Inc. & FDC Consultants Inc., 1985).
Detectable concentrations of diazinon occurred in less than 0.1%
of water samples collected from 11 Southern Ontario agricultural
watersheds during 1975-1977. The concentration was mainly below
0.01 µg/litre, the maximum value being 0.15 µg/litre (Frank et al.,
1982).
Sampling performed in 1992 by the United Kingdom National Rivers
Authority showed diazinon at >0.1 µg/litre in 74 out of 2300 fresh
water samples and at > 0.15 µg/litre in 1 out of 12 seawater samples.
5.1.3 Soil
In 1971 hay and soil samples were collected in 9 states in the
USA to determine the incidence and levels of pesticide residues in
hayfields. Residues were detected in 8% of the soil samples and 29% of
the hay samples. Diazinon was detected in four hay samples (Gowen et
al., 1976).
In 1976, soil samples from 28 farms located in six vegetable
growing areas of southwestern Ontario, Canada, contained diazinon
residues from trace amounts (< 0.02 mg/kg) to 0.29 mg/kg (Miles &
Harris, 1978b).
5.1.4 Fruit, vegetables and food
Results of supervised trials and monitoring of diazinon residues
in or on food and feed commodities have been comprehensively reviewed
and summarized (FAO/WHO, 1994a). The following examples indicate that
diazinon residues are generally low.
Ward et al. (1972) performed a study to determine the rate of
decline of diazinon residue on wheat in Texas, USA. There was a steady
decline in the amount of diazinon remaining on foliage samples after
application. Only 0.16 mg/kg and 0.31 mg/kg remained 28 days after
treatment with 0.28 and 0.56 kg a.i./ha, respectively. Harvest samples
showed that less than 0.05 mg/kg remained in either the foliage or
grain.
Between 1978 and 1986, 305 samples of apples were analysed for
residues of a wide range of pesticides used in their production.
Residues of diazinon were found occasionally. They were well below the
maximum residue limit and correlated well with the pattern of use
(Frank et al., 1989).
Between 1986 and 1988, 433 composite vegetable samples
representing 16 commodities, which were treated by various pesticides
including diazinon, were collected from farm deliveries to the
marketplace in Ontario, Canada. All samples were analysed for
insecticides and fungicides. The commodities tested included
asparagus, beans, carrots, celery, cucumbers, lettuce, onions,
peppers, potatoes, radishes, rutabagas and tomatoes. In 64% of
samples, no pesticide residues were identified (the limits of
detection ranged from 0.005 to 0.05 mg/kg). A further 22% had combined
insecticide and fungicide residues below 0.1 mg/kg. Only three samples
(0.7%) had residues that exceeded the Maximum Residue Limit (MRL).
These involved diazinon on celery. While some commodities had no
detectable residues, others had measurable residues of up to three
different pesticides. The highest levels were found on celery, lettuce
and field tomatoes (Frank et al., 1990).
Levels of diazinon permitted in the USA on human food range from
0.1 mg/kg in potatoes to 0.7 mg/kg in most leafy vegetables. During
the course of pesticide surveillance of vegetables, an unknown
analytical response in spinach extract was seen, which was
subsequently identified as diazinon metabolite (2-isopropyl-4-
methyl-pyrimidin-6-ol). Cairns et al. (1985) described an analytical
procedure adapted to confirm both diazinon and its metabolite in
spinach, at very low levels, by methane chemical ionization mass
spectrometry. The presence of this metabolite at the 1 mg/kg level
represents an order of magnitude greater than that found for diazinon
itself.
In a study of diazinon residues in prepared foods, accidentally
exposed during and following treatment, the amounts of diazinon
residues in food left in the room for 30 min after treatment ranged
from 0.02 to 0.05 mg/kg. No detectable residues of diazinon were found
in the potatoes or dinners placed in the rooms 4.5 h after treatment
and removed after 5 h. A person consuming a dinner at the highest
residue found would have ingested 0.0153 mg of diazinon. For a person
weighting 70 kg this would amount to 0.218 µg/kg (Jackson & Wright,
1975).
5.1.5 Milk
Insecticides in polyvinyl chloride pellets were included in a
commercial dairy protein supplement and fed to dairy cows at 1.4, 2.0
and 2.5 mg of diazinon/kg body mass for 2 weeks. No insecticidal
residues were found in milk samples collected at 1, 3, 7, 10 or 14
days. Even 2.5 mg/kg dosage would provide a 5-fold margin of safety
for PVC formulation-diazinon fed to cattle to control face fly larvae
in manure (Lloyd & Matthysse, 1971), and diazinon-PVC was found to
be still a highly effective larvicide if given at the dose of
0.5 mg insecticide/kg per day (Lloyd & Matthysse, 1966, 1970).
Derbyshire & Murphy (1962) reported no diazinon residues in milk
from cows fed 10 mg/kg body weight for 7 days. Robbins et al. (1957)
found only traces of radioactivity in a cow's milk 6-24 h after a
single oral dose of 32P-labelled diazinon (20 mg/kg).
5.1.6 Meat and fat
Tissue residues were determined and toxicity symptoms were noted
after lambs were sprinkled and dipped with 0.06% diazinon emulsion or
sprinkled with 1% diazinon emulsion. The only diazinon residues found
were 1.45-2.30 mg/kg in fat, 1 day after dipping in 0.06% diazinon,
with concurrent 44-47% plasma cholinesterase activity depression. Low
residues were present in blood from these sheep. Most tissues
contained no detectable diazinon at 15 or 26 days after lambs were
dipped in 0.06% diazinon, but fat contained up to 0.52 mg/kg at 15
days and 0.31 mg/kg at 26 days. Sprinkling with 1% diazinon produced
no residues in most tissues. A maximum of 23 mg/kg was found in fat.
The only clinical poisoning involved a 3-day-old lamb dipped in 0.12%
diazinon suspension. Lambs more than 1 week old were not poisoned by
0.06% diazinon nor were lambs more than 1 month old when treated by
0.25% diazinon (Matthysse et al., 1968).
Harrison et al. (1962) found 0.4 mg diazinon/kg in meat of
unshorn sheep 1 day after dipping in 0.05% diazinon emulsion. This
decreased to 0.25 mg/kg and 0.16 mg/kg at 4 and 7 days after dipping,
respectively, and there were negligible amounts 25 days after dipping.
Claborn et al. (1963) found 0.69 mg/kg in beef fat 1 day after
the last of 11 weekly spraying with 0.05% diazinon suspension. The
authors reported a rapid loss of diazinon from beef fat and the amount
of residues were negligible 14 days after spraying.
Samples obtained from retail outlets in the United Kingdom during
1984-1986 generally showed zero or low levels of diazinon residues.
Diazinon was not detected in samples of beef, imported lamb, pork or
veal, but low levels were found in United Kingdom lamb in 1984/1985
(up to 1.7 mg/kg) and 1985/1986 (up to 0.1 mg/kg). Samples of fat
taken in 1986 were analysed and, out of 274, 19% contained diazinon.
In 1987, however, out of 280 samples analysed, 7% contained diazinon
and in four of them residues exceeded the Codex MRL of 0.7 mg/kg fat.
Diazinon was not detected in butter, milk or cheese (MAFF, 1989).
Various pesticides and pollutants were examined in poultry meat
from Israel. The levels of these, which included diazinon in broilers,
turkeys and geese, were said to be extremely low and below the USA
tolerance levels (Kathein, 1986)
5.2 General population exposure
The primary exposure to the general population will be through
intermittent dietary exposure and inhalation exposure. Exposure via
water is negligible. Total-diet studies commenced in the United
Kingdom in 1966. In the second survey (1970-1971) and in the latest
survey (1985-1988), diazinon residues were not detected (Egan &
Weston, 1977; MAFF, 1982, 1986, 1989). Findings similar to those in
the United Kingdom were also made in the USA. Toddler total diets have
also been the subject of investigation in the USA. Diets collected in
ten American cities between 1978 and 1979 were examined. The
components were drinking-water, whole milk, other dairy products and
dairy substitutes, meat/fish/poultry, grain cereals, potatoes,
vegetables, fruit juices, oils and fats, sugars and beverages
(Gartrell et al., 1985a). A similar exercise in the years 1980-1982
was conducted in 13 American cities. The results were similar to those
obtained in 1978-1979, with intake of diazinon being low (Gartrell et
al., 1985a,b).
A total-diet study in New Zealand was performed at 3-monthly
intervals in the period 1974-1975. Of 116 samples analysed, 82 (71%)
had no detectable residues of diazinon. Intakes were well below the
Codex MRLs (Dick et al., 1978).
The overwhelming evidence from residue and total-diet studies
suggests that residues of diazinon are generally within the acceptable
levels set by the Codex Alimentarius Commission. The results suggest
that the compounds are rapidly broken down, whether on plants or in
animals, further reducing the risks to humans (IPCS, 1986).
During a 5-year study period (1981-1986), the US Food and Drug
Administration analysed nearly 20 000 domestic and imported samples of
food and feed commodities for pesticide residues. The results showed
that 29 out of 6391 domestic agricultural commodities and 35 out of
12 044 imported agricultural commodities had diazinon levels greater
than 0.05 mg/kg (Hundley et al., 1988).
Diethyl phosphate (DEP), an organophosphate metabolite, was found
in the urine of symptomatic residents who resided in a household that
had been sprayed with diazinon 4.5 months earlier. Pre- and
post-decontamination data with regard to symptoms and to DEP,
cholinesterase, and surface and air levels underscore the utility of
alkyl phosphate metabolites for monitoring exposure. The data also
emphasize the efficacy of clean-up measures when baseline data are not
available to determine if "within-normal" cholinesterase levels are,
in fact, depressed (Richter et al., 1992).
5.3 Occupational exposure
An occupational exposure study was conducted for a firm employing
22 pest control operators (PCOs) exposed to three organophosphorus
insecticides including diazinon. The 8-h exposure levels were less
than 131.0 µg/m3. Urine samples (24-h) were analysed for alkyl
phosphates and showed the presence of metabolites for these three
insecticides. The effect of this exposure was reflected by a
statistically significant inhibition of plasma cholinesterase activity
among the PCOs, but physical examinations detected no apparent toxic
effects (Hayes et al., 1980).
A behavioural evaluation of pest control workers with short-term
(mean 39 days) low-level exposure to diazinon was conducted in 1985
during the course of a pest control program in California (see section
8.2.2). The diazinon metabolite diethylthiophosphate (DETP) was
measured in pre- and post-shift urine samples and the full-shift
exposure to diazinon was quantified for 19 subjects using personal
air monitoring and passive badges. The median diazinon exposure was
2.1 mg/day (Maizlish et al., 1987).
An investigation was conducted to determine worker exposure to
airborne pesticides during tree and ornamental shrub applications
using hand-held equipment during an entire work shift. Employee
exposure data were collected for 3 consecutive years. The sampling was
performed during the late spring, summer and early autumn when insect
and disease activity was most prevalent. Sampling was conducted at 23
locations. Those applying these chemicals sustained low-level exposure
to acephate, benomyl, carbaryl, chlorothalonil, diazinon and dicofol.
As pesticide label instructions for mixing and applying pesticides
were strictly followed, the tree and ornamental shrub applicators were
able to keep inhalation exposures below the levels recommended by OSHA
and NIOSH. Of the 74 exposures monitored, 67% were below the detection
limit (0.001 mg/m3), while others were 0.001-0.040 mg/m3. This
observation supports the correctness of not including specific
respiratory protection measures on pesticide label directions for
mixing, loading and applying these pesticides (Leonard & Yeary 1990).
Dermal, respiratory and urine measurements were made on workers
applying granular diazinon pesticide formulation. In all, 15 workers
and four control subjects were monitored. The workers applied the
compound in yards and small pastures using hand equipment comparable
to that used in a home environment. Respiratory air samples, ethanol
hand rinse samples, clothing patch samples and urine samples were
collected. The diazinon exposures were correlated with job category,
application duration, application equipment and protective clothing.
The best determinants of diazinon exposure were the job categories and
the use of the belly grinder type of spreader. The rank of exposure
magnitude, from highest to lowest, was the crew using the belly
grinder, the crew not using the belly grinder, the crew chief and the
supervisor. The mean daily dermal and respiratory diazinon exposures
for these four job categories ranged from 0.6 to 11 mg, 0.1 to 1.8 mg,
0.1 to 0.25 mg, and 0.03 to 0.07 mg, respectively. The amount of
urinary diethylthiophosphate increased during the day for all job
levels, but showed variable recovery (Weisskop et al., 1988).
6. KINETICS AND METABOLISM
6.1 Absorption, distribution and excretion
6.1.1 Oral administration
6.1.1.1 Rats
Four male and 2 female Wistar rats were treated with single
oral doses of 0.8 mg [pyrimidine-14C]-diazinon (specific activity
4.0 µCi/mg). An additional group of 4 males received [ethoxy-14C]-
diazinon (3.2 µCi/mg) at the same dose level. During the observation
period of 168 h, both labelled parts of the molecule were excreted
almost completely, 65.4-80.0% of the administered radioactivity being
detected in urine, 16.0-23.5% in the faeces and, with the ethyl-label,
5.6% in the expired air (total recovery 90.2-98.3%). No radioactive
CO2 was detected with the pyrimidine label. The half-life times of
excretion were 7 h with the ethyl label and 12 h for both sexes
treated with the pyrimidine-labelled material. Daily oral
administration of 0.1 mg [pyramidine-14C]-diazinon to male rats for
10 consecutive days resulted in no accumulation of the radioactivity
in any organ investigated (oesophagus, stomach, intestines, liver,
spleen, pancreas, kidneys, lungs, testes, muscles, fat). Six hours
after the last administration, the highest residues were detected in
the muscles (0.77% of the totally applied dose), caecum (0.76%) and
small intestine (0.65%). The residues were below the detectable limit
48 h after the cessation of the treatment (Mücke et al., 1970).
Sprague Dawley rats received [pyrimidine-14C]-diazinon at single
oral doses of 10 mg/kg (specific activity 30.3 µCi/mg) or 100 mg/kg
(specific activity 9.7 µCi/mg). A third group was treated with daily
oral doses of 10 mg/kg technical diazinon (87.7% pure) for 14
consecutive days, followed by a single treatment at the same dose
level with the 14C-labelled compound. The disposition of the
administered 14C was observed for a 7-day period before the animals
were killed and the tissues removed for analysis. The average recovery
of the radioactivity was 99.2%. Elimination of diazinon equivalents
was rapid. In the low-dose group, males and females eliminated 93 and
86%, respectively, of the administered radioactivity in the urine
within 24 h. Faecal elimination amounted to 1.6 and 1.1%,
respectively, in the same time period. In the high-dose group, the
respective values were slightly lower and indicated that the
elimination was more rapid in males (90.8% in urine and 2.2% in
faeces) than in females (58.2% in urine and 0.87% in faeces). The
pre-conditioning of the rats had no influence on absorption and
elimination. Seven days after the administration of the [pyrimidine
14C]-diazinon, the residual radio-activity was generally low. Among
the tissues examined (heart, lung, spleen, kidney, liver, fat, testes,
ovaries, uterus, muscle, brain, blood plasma, blood cells, bone), the
residual radioactivity amounted to approximately 0.01 mg/kg diazinon
equivalents in the low-dose group; only fat (0.02 mg/kg), blood cells
(0.05 mg/kg) and bone (<0.017 mg/kg) contained higher amounts of
radioactivity. In the high-dose group, the residual radioactivity was
8-10 times higher. Pretreatment with technical diazinon for 14 days
led to residues similar to those observed in the low-dose group
(Craine 1989a,b).
6.1.1.2 Guinea-pigs
Male guinea-pigs treated orally with 45 mg/kg [32P]-diazinon
(specific activity 117-197 cpm/mg) in peanut oil, the tissue
distribution was determined at 2, 4, 8 and 16 h after treatment and
the excretion of 32P was investigated over an 8-day period. Following
oral administration, the compound was rapidly absorbed as shown by a
sharp decrease of activity in the stomach and low levels found in the
small intestine. Within 16 h, 46.6% of the administered radioactivity
was eliminated in the urine and 0.34% appeared in the faeces. The
caecum showed a gradual increase of radioactivity, 13-36% of the
administered dose accumulating in the caecum over 16 h after the
administration. Irrespective of this accumulation, within 48 h after
dosing, 80% of the administered radioactivity was eliminated in the
urine while only 8% was eliminated in the faeces (Kaplanis et al.,
1962).
6.1.1.3 Dogs
Two female Beagle dogs were intravenously dosed with 0.2 mg/kg
[ethoxy-14C]-diazinon (specific activity 3.4 µCi/mg) in 0.7 ml
ethanol. Blood samples were drawn at times ranging from 5 min to 7 h
after the injection. The decline of the radioactivity in the blood
was biphasic with a slower second phase. The half-life of elimination
from blood for this second phase was calculated to be 363 min.
Approximately 58% of the administered radioactivity was recovered in
the urine within 24 h after the administration. Another two female
beagle dogs were orally dosed by capsule with 4.0 mg/kg [ethoxy-14C]
diazinon in ethanol. Approximately 85% of the administered
radioactivity was recovered within 24 h after oral administration,
with 53% of it occurring in urine (Iverson et al., 1975).
6.1.1.4 Goats
Two lactating goats were orally treated with [pyrimidine-14C]-
diazinon (specific activity 9.7 µCi/mg) in gelatin capsules for four
consecutive days at a dose level of 4.5 mg/kg per day, corresponding
to a dietary exposure of 100 mg/kg of feed. During the observation
period, in average 64.1% of the administered radioactivity was
excreted with urine, 10.4% with the faeces and 0.31% with the milk.
A plateau of radioactivity in the milk was reached after 3 days of
dosing at a mean level of 0.46 mg/kg diazinon equivalent. At
sacrifice, radioactivity in the blood accounted for 0.2% and the
tissues examined accumulated 0.92% of the administered dose. The
highest residual radioactivity was detected in the kidney (2.0 mg/kg)
and the liver (1.2 mg/kg). The other tissues examined contained
0.23-0.3 mg/kg diazinon equivalents (Simoneaux 1988a,b; Pickles &
Seim, 1988).
6.1.1.5 Cow
A lactating Hereford cow (body weight 268 kg) was orally treated
with a gelatin capsule containing 20 mg/kg 32P-diazinon (specific
activity 518 cpm/µg). Urine and faeces were collected during 36 h
after treatment and further samples were investigated until the study
was terminated after 168 h. In addition, milk and blood samples were
investigated. Within 36 h, approximately 74% of the administered
radioactivity was excreted with the urine, 6.5% appeared in the faeces
and 0.08% was found in the milk. A peak concentration of 2.27 mg/kg
diazinon equivalents was reached 18 h after the administration
(Robbins et al., 1957).
6.1.1.6 Hens
Four laying Leghorn hens were treated with 2-14C-diazinon
(specific activity 30.3 µCi/mg) in gelatine capsules for seven
consecutive days at daily doses of 1.7 mg/kg body weight,
corresponding to a dietary exposure of 25 mg/kg in feed. Excreta and
eggs were collected and, approximately 24 h after the final dose, the
animals were killed and tissue samples of liver, kidney, blood, lean
meat, skin and attached fat, and peritoneal fat were examined.
Elimination of most of the administered radioactivity occurred via the
excreta, with 78.6% of the total dose being excreted during the study
period. Approximately 0.1% of the radioactivity was found in tissues
and blood, less than 0.01% appeared in the egg yolks and 0.07% was
detected in the egg whites. The residual radioactivity in the
tissues amounted to 0.148 mg/kg diazinon equivalents in the kidney,
0.137 mg/kg in blood, 0.11 mg/kg in the liver and 0.01-0.025 mg/kg in
the other tissues examined. The residues in the egg yolks ranged from
0.006 mg/kg diazinon equivalents to 0.065 mg/kg while those in the egg
whites ranged from 0.038 mg/kg to 0.066 mg/kg. On a whole egg basis, a
plateau concentration of 0.047 mg/kg was reached on day 4 of treatment
(Simoneaux 1988c,d; Burgener & Seim, 1988).
6.1.2 Dermal application
6.1.2.1 Rats
The percutaneous absorption of diazinon was investigated in male
and female Sprague Dawley rats dermally exposed to 1 mg/kg (specific
activity 25.2 µCi/mg) and 10 mg/kg (specific activity 2.62 µCi/mg) of
[pyrimidine-14C-diazinon] dissolved in tetrahydrofuran. The dermal
absorption, excretion and tissue residues were determined after 0, 2,
8, 24, 48, 72 and 144 h. At each time point, four rats per sex and
dose group were used. The total recoveries for the balance data
averaged 96.3-101.5%. Calculated t50 absorption rates (i.e. the
amount of time required for 50% of the administered dose to be
absorbed into or penetrate through the skin) in males and females were
11.8 and 5.2 h, respectively, at the low-dose level of 1 mg/kg. At
10 mg/kg the respective t50 absorption rates were 10.2 and 5.3 h,
respectively, indicating that dermal absorption was more rapid in
females and was dose-dependent. The urine was the major route of
excretion in both sexes at both dose levels, 65-78% of the radiolabel
being excreted within 72 h. Times for 50% excretion in males and
females dosed at 1 mg/kg were 28.1 and 26.8 h, respectively. In the
high-dose groups the times for 50% excretion were 24.1 and 20.3 h in
males and females, respectively. The residual radioactivity in tissues
reached a maximum at 8 h after the administration in both dose groups
(plasma, red blood cells, fat, brain, muscle, lung, heart, spleen,
kidney, liver, stomach, small and large intestines, gonads, skin wash
and dissolved skin were assayed). In the low-dose group of males after
8 h, highest values were found in stomach (0.36 mg/kg diazinon
equivalents), small intestines (0.16 mg/kg), kidney (0.15 mg/kg),
liver (0.1 mg/kg) and skin (3.9 mg/kg in the skin wash and 0.86 mg/kg
in the dissolved skin). Reflecting their absorption rate, the females
of the low-dose group showed slightly higher tissue levels and a lower
residual radioactivity in the skin wash. After 144 h, residues were
down to the limit of quantification in most tissues, in both dose
groups and in both sexes (Ballantine, 1984).
6.1.2.2 Sheep
Two sheep were dermally treated with [pyrimidine-14C-diazinon]
(specific activity: 3.7 µCi/mg) dissolved in acetone for three
consecutive days. In order to mimic an extreme maximum exposure in a
dermal treatment of 40 mg/kg, 2270 mg 14C-diazinon was applied daily
to a shaved area of the back that constituted approximately 10% of the
animal's surface area. The area of application was left uncovered. Six
hours after the last administration the animals were killed and heart,
liver, kidney, back fat and leg muscle were analysed. The tissue
extractability was greater than 90% for all tissues. The highest
average residues were detected in kidney (9.4 mg/kg diazinon
equivalents) and back fat (7.3 mg/kg), while levels in heart, liver
and leg muscle amounted to 4-4.4 mg/kg (Capps, 1990; Pickles, 1990).
6.1.2.3 Humans
The dermal absorption of diazinon in humans is much less than in
rats. Six volunteers were dermally treated with [pyrimidine-14C]-
diazinon on the ventral forearm or the abdomen. The test material was
administered in acetone solution (2 µg/cm2) or dissolved in lanoline
wool grease (1.47 µg/cm2) over a 10-cm2 area of the skin without
occlusion. After 24 h, the test substance remaining on the site of
administration was washed off and the renal elimination followed for
seven days. Independent of the vehicle and the site of administration,
only 3-4% of the dose applied was percutaneously absorbed (Wester et
al., 1993).
6.1.3 Other routes
6.1.3.1 Intraperitoneal administration
The tissue distribution of diazinon and the inhibition of
cholinesterase (ChE) activities in plasma and erythrocytes were
investigated using male rats that received a single intraperitoneal
dose of diazinon (100 mg/kg body weight) in olive oil. The blood
diazinon level was estimated to reach a maximum at 1-2 h after
intraperitoneal administration. It was demonstrated that the diazinon
residue levels were highest in the kidney, when comparing the
distribution of diazinon among liver, kidney and brain in the animals
after dosing. Erythrocyte and plasma ChE activities were inhibited
rapidly, but ChE inhibition was greater in the erythrocytes than in
plasma (Tomokuni & Hasegawa, 1985).
The tissue distribution of diazinon and the inhibition of ChE
activities in plasma, erythrocyte and brain was investigated using
male rats and mice that received a single intraperitoneal (i.p.) dose
of diazinon (20 or 100 mg/kg body weight) in olive oil. The blood
diazinon level was estimated to reach a maximum 1-2 h after the i.p.
administration. It was demonstrated that the diazinon residue levels
were highest in the kidney, when comparing the distribution of
diazinon among liver, kidney and brain in the animals after dosing.
The ChE inhibition by diazinon exposure was greater in the plasma than
in the erythrocytes for male mice, while its inhibition was greater in
the erythrocytes for male rats. Brain ChE activity was also inhibited
markedly in the mice after dosing (Tomokuni et al., 1985).
6.1.3.2 Subcutaneous administration
Male guinea-pigs were subcutaneously treated with 45 mg/kg
32P-labelled diazinon (specific activity -117-197 cpm/µg) in peanut
oil. The tissue distribution was determined 2, 4, 8 and 16 h after
treatment, and excretion of 32P was investigated over an 8-day
period. Following subcutaneous administration, urinary elimination
amounted to 20% of the administered dose after 16 h. The levels of
radioactivity found in the gastrointestinal tract were low apart from
the caecum, which accumulated up to 5.5% of the administered dose over
16 h. After 48 h, urinary elimination amounted to about 60%, while
only trace amounts were eliminated with the faeces (Kaplanis et al.,
1962).
6.1.3.3 Intravenous administration
Four female Rhesus monkeys were dosed intravenously with 2.1 µCi
(31.8 µg) [pyrimidine-14C]-diazinon dissolved in propylene glycol.
Within 7 days, average values of 56 and 23% of the dose were
eliminated in urine and faeces, respectively (Wester et al., 1993).
6.2 Metabolism
The metabolic fate of diazinon was studied with different modes
of administration using unlabelled and radiolabelled diazinon in
various species including rat, mouse, guinea-pig, dog, sheep, goat,
cow and chicken. Additional in vitro experiments were conducted
using tissue slices or cell fractions. A comparative summary of the
results available was provided by Hagenbuch & Mücke (1985). In all
species tested, diazinon was rapidly and almost completely absorbed
from the gastrointestinal tract. It was also absorbed from the skin.
The main metabolic pathways of degradation of diazinon are:
a) Cleavage of the ester bond of diazinon or diazinoxon leading to
the hydroxypyrimidine derivatives.
b) Transformation of P-S moiety to the P-O derivative, leading to
the active metabolite, diazoxon.
c) Oxidation of isopropyl substituent leading to the corresponding
tertiary and primary alcohol derivatives.
d) Oxidation of the methyl substituent leading to the corresponding
alcohol.
e) Glutathione-mediated cleavage of the ester bond leading to a
glutathione conjugate.
The hydrolytic and oxidative cleavage of the phosphorus ester
bond, leading directly or via diazoxon to the pyrimidinyl derivative,
play the most prominent role in the metabolism of diazinon.
Glutathione conjugation appears to be of small importance. Metabolites
maintaining the phosphorus ester bond are of transient nature and are
only observed in minor quantities.
The general metabolic pathways of diazinon in mammals are given
in Fig. 1.
The metabolites formed, i.e. diethylphosphoric acid,
diethylthiophosphoric acid and the derivatives of pyrimidinyl ring,
are eliminated mainly via the kidneys. Only minimal quantities of the
metabolites were detected in milk and eggs.
6.2.1 In vivo metabolic transformations
6.2.1.1 Mice
When male ICR mice (number not stated) were treated orally with
diazinon or [pyrimidine14C] diazinon at 50 or 75 mg/kg body weight,
one half of the high-dose animals died and the rest showed symptoms
(sweating, crouching) (Miyazaki et al., 1970; Sekine, 1972). At the
low dose, no signs of toxicity were observed. Metabolism and excretion
occurred rapidly, and the metabolites diazoxon, O, O-diethyl-
O-[2-(alpha-hydroxyisopropyl)-4-methyl)-6-pyrimidinyl]
phosphorothioate, and O, O-diethyl- O-(2-(2-propenyl)-4-methyl-6-
pyrimidinyl) phosphorothioate were found in the urine 1 h after
treatment. Most of the metabolites were found in urine 6 h after
treatment, but metabolism was not identical in the two dose groups. In
the low-dose group O, O-diethyl- O-(2-isopropyl-4-hydroxymethyl-6-
pyrimidinyl) phosphorothioate and O, O-diethyl- O-(2-isopropyl-4-
formyl-6-pyrimidinyl) phosphorothioate were found, but this was not
observed in the high-dose group. In the high-dose, but not the low-
dose group O, O-diethyl- O-(2-(a-hydroxyethyl)-4-methyl-6-
pyrimidinyl) phosphorothioate was found.
The metabolism of [pyrimidine-14C]-diazinon and [ethoxy-14C]-
diazinon was investigated by Mücke et al. (1970). Four metabolite
fractions were found in urine and faeces, three metabolites
representing approximately 70% of the total radioactivity applied.
Hydrolysis of the ester bond yielded 2-isopropyl-4-methyl-6-
hydroxypyrimidine (22.5% of the applied radioactivity in urine);
oxidation at the primary carbon atom produced 9% of the applied
radioactivity in urine, while oxidation at the tertiary carbon atom of
the isopropyl side chain produced 22%. In addition, trace amounts of
unchanged diazinon were detected in faeces. No cleavage of the
pyrimidine ring with subsequent oxidation of the fragments to CO2
took place (Mücke et al., 1970).
6.2.1.2 Rats
A study by Capps (1989) investigated the diazinon metabolites in
male and female rats orally treated with single doses of 10 and
100 mg/kg [pyrimidine 14C]-diazinon and in rats preconditioned with
14 daily treatments at 10 mg/kg before the final administration of
radiolabelled compound. The metabolite pattern was similar in the
urine and faeces of the rats from all dose groups and from both sexes.
The major urinary metabolites were identified as 2-isopropyl-6-
methyl-4(1 H)- pyrimidinone (average 38.2% of the totally applied
dose), 2-(alpha-hydroxyisopropyl)-6-methyl-4(1 H)-pyrimidinone
(17.3%) and 2-(beta-hydroxyisopropyl-6-methyl-4(1 H)-pyrimidinone
(9.7%). Six unknown aqueous components accounted for an average of
14.9% of the administered dose, and trace amounts of unchanged
diazinon (0.11%), diazoxon (0.14%) and the hydroxy-isopropyl
derivative of diazinon (0.12%) were also detected. The identity of the
metabolites was confirmed by gas chromatography and mass spectrometry
(GC/MS) with synthetic standards.
6.2.1.3 Dogs
The urinary metabolites of Beagle dogs were characterized after
oral administration of 4.0 mg/kg body weight 14C-ring-labelled
diazinon. The metabolite 2-isopropyl-4-methyl-6-hydroxypyrimidine
accounted for 10% of the applied radioactivity in the urine and the
tertiary hydroxy-isopropyl derivative of diazinon represented 23%
(Iverson et al., 1975).
6.2.1.4 Sheep
When two sheep were dermally treated with [pyramidine-14C]-
diazinon, radiolabelled residues were detected in all tissues examined
(heart, liver, kidney, back fat and leg muscle). Unmetabolized
diazinon was the only significant residue in fat, and was a major
residue in heart and leg muscle. The major metabolites in urine and
all tissues except fat were 2-isopropyl-6-methyl-4(1 H)-pyrimidinone
(urine, 10% of the administered radioactivity; liver, 18%; kidney,
23%) and 2-(alpha- hydroxyisopropyl)-6-methyl-4(1 H)-pyrimidinone
(urine, 22.7%; liver, 10%; kidney, 28%), which were also present in
the form of glucuronide conjugates. The identity of the metabolites
was confirmed by GC/MS with synthetic standards. In addition, several
unidentified polar (urine, 18.6%) and minor amounts of non-polar
(urine, 4.0%) metabolites were detected (Capps, 1990).
6.2.1.5 Goats
Two lactating goats were orally treated with [pyrimidine-14C]-
diazinon in gelatin capsules for four consecutive days. Similarly to
sheep, in urine and faeces the metabolites 2-isopropyl-6-methyl-
4(1 H)-pyrimidinone (urine, 4.5% of the totally administered radio-
activity; faeces, 2.6%) and 2-(alpha-hydroxyisopropyl)-6-methyl-
4(1 H)-pyrimidinone (urine, 12.5%; faeces, 1.7%) were identified.
Approximately 48.6% of the urinary radioactivity consisted of unknown
water-soluble compounds. Characterization of selected tissues showed
the presence of mainly the above-mentioned metabolites. Unchanged
diazinon, its hydroxy-isopropyl derivative and diazoxon accounted for
less than 10% of the radioactivity detected in these tissues.
Metabolites in fat consisted primarily of unchanged diazinon (66%),
its hydroxy-isopropyl derivative (12.5%) and diazoxon (3%). The major
metabolites in the milk were 2-isopropyl-6-methyl-4(1 H)-
pyrimidinone (39.3% of the residual radioactivity) and
2-(alpha-hydroxyisopropyl)-6-methyl-4(1 H)-pirimidinone (37.3%).
Substantial portions of the polar metabolites in urine, faeces and
tissues were glucuronide conjugates. The identity of the metabolites
was confirmed by GC/MS with synthetic standards (Simoneaux,
1988a,b,e).
6.2.1.6 Hens
Four laying Leghorn hens were treated with [pyrimidine-14C]-
diazinon in gelatin capsules for seven consecutive days at daily doses
of 2.75 mg/kg day. The main metabolites detected in the excreta were
unchanged diazinon (14.9% of the extractable radio-activity),
2-isopropyl-6-methyl-4(1 H)-pyrimidinone (5.9%), 2-(alpha-hydroxy-
isopropyl)-6-methyl-4(1 H)-pyrimidinone (10.8%) and 2-(beta-
hydroxyisopropyl)-6-methyl-4(1 H)-pyrimidinone (7.2%). Approximately
25% of the radioactivity in the excreta consisted of unknown
water-soluble compounds. The residues in tissues primarily consisted
of 2-isopropyl-6-methyl-4(1 H)-pyrimidinone (0.6-2.6% of the residual
radioactivity), 2-(alpha-hydroxyisopropyl)-6-methyl-4(1 H)-
pyrimidinone (3.1-6.5%) and 2-(beta-hydroxyisopropyl)-6-methyl-
4(1 H)- pyrimidinone (2.0-5.7%). Unchanged diazinon was detected
primarily in the peritoneal fat (2% of residues). In the eggs,
primarily 2-isopropyl-6-methyl-4(1 H)-pyrimidinone (yolk, 11.1% of
the residual radioactivity; white, 9.4%), 2-(alpha-hydroxyisopropyl)-
6-methyl-4(1 H)-pyrimidinone (yolk, 18.6%; white, 33.3%) and
2-(beta-hydroxyisopropyl)-6-methyl-4(1 H)-pyrimidinone (yolk, 7.0%;
white, 35.3%) were detected. As in goats, a substantial portion of the
polar metabolites in tissues, eggs and excreta were glucuronide
conjugates. The identity of the metabolites was confirmed by GC/MS
with synthetic standards (Simoneaux, 1988c,e; Simoneaux, 1989).
More information on kinetics and metabolism in other species is
given in chapter 9.
6.2.2 In vitro metabolic transformations
The metabolism of [ethoxy-14C]-diazinon and diazoxon was studied
in vitro using rat liver cell fractions. It was shown that the
degradation by diazinon is catalysed by a microsomal enzyme that
requires NADPH and oxygen, and is inhibited by carbon monoxide. It is
presumably the cytochrome P-450 oxidase system. Diazoxon was shown to
be degraded by enzymes located in the nuclear, mitochondrial,
microsomal and soluble fractions of the liver. The microsomal enzymes
were the most active and were not dependent on NADPH. Reduced
gluthation had little effect. With diazinon, products of the reactions
were diethylphosphorothioic acid and diethylphosphoric acid. Diazoxon
was degraded to diethylphosphoric acid (Yang et al., 1969, 1971;
Nakatsugawa et al., 1969). These results were confirmed by independent
experiments (Dahm, 1970). The oxidation of diazinon was investigated
by using microsomal preparations from rat liver. The major metabolic
products of diazinon were hydroxydiazinon, diazoxon and
hydroxydiazoxon, which are biologically active, and additional
inactive products such as diethylphosphorothioic acid,
diethylphosophoric acid and derivatives of the pyrimidyl moiety. It
was demonstrated that desulfuration, hydroxylation of the ring alkyl
side-chain and cleavage of the aryl phosphate bond may occur,
depending on the presence of NADPH or NADH. EDTA stimulated the
overall metabolism of diazinon (Shishido et al., 1972a).
The enzymatic hydrolysis of diazoxon was investigated using rat
tissue homogenates. The hydrolytic activity of the tissues decreased
in the order liver>blood>lung>heart>kidney>brain. In the liver,
the hydrolytic activity was localized in microsomal preparations.
Diethyl phosphoric acid and 2-isopropyl-4-methyl-6-hydroxypyrimidine
were identified as the products. The reactions were inhibited by EDTA,
heavy and rare earth metal ions, and sulfhydryl reagents (L-cysteine,
2-mercaptoethanol, thioglycolic acid), while calcium ions activated
the hydrolysis (Shishido & Fukami, 1972).
Liver homogenates were prepared from male mice (North Carolina
Department of Health strain) and incubated for 1 h with either
14C-diazinon or 14C-diazoxon. Inhibition of metabolism was
studied by co-incubation with piperonyl butoxide, NIA 16824 or
1-(2-isopropylphenyl) imidazole. Diazoxon formation from diazinon
(thiophosphate to phosphate conversion) was inhibited by 45 to 60% by
the inhibitors studied. All the inhibitors also reduced oxidative
dearylation of diazinon to diethyl phosphoric and diethyl
phosphorothioic acids (Smith et al., 1974).
Conjugation with glutathione forms the third enzymatic mechanism
of the diazinon metabolism in rat tissue preparations (liver, heart,
brain, lung, kidney and blood were investigated). The highest activity
(14-89 times as high as in other tissues) for this reaction was
localized in the cytoplasmatic fractions of the liver. The reaction
products were identified as diethyl phosphorothioic acid and
S-(2-isopropyl-4-methyl-6-hydroxypyrimidinyl) glutathione, which were
formed by conjugation and simultaneous cleavage of the phosphate ester
bond. The enzymatic activity was increased by the addition of
glutathione-SH, and was inhibited by various sulfhydryl reagents,
oxidized glutathione and some chelating agents ( o-phenanthroline,
8-hydroxyquinoline) (Shishido et al., 1972b).
6.3 Metabolic aspects of diazinon toxicity
Diazinon was incubated with liver microsomes and liver slices
from sheep, cow, pig, guinea-pig, rat, turkey, chicken and ducks.
Hydroxydiazinon, isohydroxydiazinon, dehydrodiazinon, their oxons
and diazoxon were identified and determined quantitatively or
semi-quantitatively. It was shown that yields and rates of production
of the metabolites varied greatly between the species. The production
of the oxon was not generally correlated with susceptibility to
diazinon poisoning, although it was lowest in the least susceptible
animal, the sheep. The highly susceptible avian species (acute oral
LD50 of around 2-15 mg/kg) do not produce higher rates of oxons than
rat or pig (acute oral LD50 around 300-600 mg/kg). However, the
mammalian blood hydrolyses diazoxon rapidly, whereas the avian species
have virtually no hydrolytic activity. It was concluded that
extrahepatic metabolism of diazinon, in particular the hydrolysis of
diazoxon in the blood, appears to be the main factor affecting
susceptibility to diazinon poisoning. In mammals the extrahepatic
metabolism of diazinon is more important toxicologically than the
metabolism in the liver, while the liver is probably the most
important site of metabolism in avian species (Machin et al., 1975).
Recently, the hydrolytic metabolism of diazinon by plasma was
investigated in 92 individuals of Hispanic origin (Davies et al.,
1996). Diazoxon is hydrolysed by the enzyme paraoxonase (PON1),
leading to the formation of 2-isopropyl-4-methyl-6-hydroxypyrimidine
and diethylphosphate. An important observation of this study was that
the effect of the PON1 polymorphism for diazoxon hydrolysis relative
to paraoxon hydrolysis was reversed. Thus, RR individuals (Arg192
homozygotes) who displayed high paraoxonase activity had lower
diazonoxonase activity (mean = 7948 U/litre) than QQ homozygotes
(12 318 U/litre).
7. EFFECTS ON LABORATORY MAMMALS AND IN VITRO TEST SYSTEMS
7.1 Single exposure
7.1.1 Oral
Improvements since 1979 in the manufacturing of diazinon have
significantly reduced the content of highly toxic by-products, in
particular tetraethyl-pyrophosphate (TEPP). As a result of these
stepwise improvements, the acute oral LD50 of technical grade
diazinon increased to values around 1000 mg/kg (Piccirillo, 1978;
Bathe & Gfeller, 1980; Schoch & Gfeller, 1985; Kuhn, 1989a). The most
recent study resulted in an oral LD50 in rats of 1250 mg/kg. The