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    PHOSMET (addendum)      JMPR 1998

    First draft prepared by
    T.C. Marrs
    Department of Health
    London, United Kingdom


         Explanation
         Evaluation for acceptable daily intake 
              Delayed neuropathy 
              Unscheduled DNA synthesis  in vivo
         Comments 
         Toxicological evaluation 
         References 


    Explanation

         Phosmet was evaluated toxicologically by the Joint Meeting in
    1978 (Annex 1, reference 30), when a temporary ADI of 0-0.005 mg/kg bw
    was allocated. It was re-evaluated in 1979 (Annex 1, reference 32)
    when additional data on teratogenicity became available, and an ADI of
    0-0.02 mg/kg bw was established. Further data were made available to
    the 1994 JMPR (Annex 1, reference 73), when an ADI of 0-0.01 mg/kg bw
    was allocated on the basis of the NOAEL in a multigeneration study of
    reproductive toxicity in rats, with a 100-fold safety factor. At that
    meeting, further information was requested, namely a further long-term
    study in dogs, studies on DNA binding  in vivo, and a study of the
    potential of phosmet to cause delayed polyneuropathy in hens at an
    appropriately high dose, with an estimate of neuropathy target
    esterase. The last study was supplied but not the requested study of
    repeated dosing in dogs; moreover, a study of unscheduled DNA
    synthesis in rat liver  in vivo was submitted rather than a study of
    DNA binding  in vivo. The new studies are reviewed below.

    Evaluation for Acceptable Daily Intake

    1.  Delayed neuropathy

         A study of the ability of phosmet to cause delayed polyneuropathy
    in Lohmann Brown hens  (Gallus gallus domesticus) was conducted with
    phosmet of a purity of 97.4%. In a preliminary study, the LD50 of
    phosmet given orally was determined in seven groups of 10 birds. It
    was estimated to be 580 mg/kg bw (95% confidence interval, 410-770). A
    dose of 600 mg/kg bw was therefore given to 24 birds; 12 further birds
    received the vehicle (corn oil), and a further 12 were given
    tri- ortho-cresyl phosphate at 1 g/kg bw as positive controls. All
    birds were injected subcutaneously with atropine sulfate at 20 mg/kg
    bw just before treatment, and further injections were given to birds
    that showed severe clinical signs. The birds were observed for adverse
    clinical signs, ataxia, and effects on body weight. After 48 h, three
    birds from the groups receiving phosmet, the vehicle or the positive

    control were killed, and brain acetylcholinesterase and brain and
    spinal cord neuropathy target esterase activities were estimated.
    After 21 days, six birds from each group were sacrificed and fixed by
    perfusion, and the head, spinal column, and dissected sciatic and
    tibial nerves were taken and stored in 10% buffered formalin. Sections
    of fore-, mid-, and hindbrain; upper and lower cervical, mid-thoracic,
    and lumbo-sacral spinal cord; and proximal and distal sciatic and
    tibial nerves were examined histopathologically. 

         No ataxia was observed in the birds treated with phosmet, while
    8/12 birds treated with tri- ortho-cresyl phosphate developed
    clinical ataxia, commencing at 11-18 days. Brain acetylcholinesterase
    activity was reduced in birds given phosmet to 37% of the value in
    concurrent vehicle controls, but the neuropathy target esterase
    activity in both brain and spinal cord was similar to that of
    controls. In contrast, in birds given tri- ortho-cresyl phosphate,
    the acetylcholinesterase activity was similar to that of vehicle
    controls, whereas the neuropathy target esterase activity was markedly
    inhibited, being 9.6% of the value for concurrent controls in brain
    and 20% that of concurrent controls in spinal cord. Vehicle controls
    gained weight normally, while those treated with tri- ortho-cresyl
    phosphate lost weight. With phosmet, initial weight loss was followed
    by recovery. Histopathological examination showed no evidence of the
    characteristic changes of delayed neuropathy in the phosmet-treated or
    vehicle control hens, whereas birds that had received
    tri- ortho-cresyl phosphate showed minimal axonal degeneration in the
    cerebellum and minimal or moderate axonal degeneration at one or more
    levels of the spinal cord and in some peripheral nerve sections
    (Johnson, 1997).

    2.  Unscheduled DNA synthesis  in vivo

         The ability of phosmet (purity, 96.4%) to induce unscheduled DNA
    synthesis in male Alpk:APfSD rats was determined by an
    autoradiographic technique. Five rats received phosmet orally at a
    dose of 32 (two rats) or 50 mg/kg bw (three rats) in corn oil, the
    higher dose being the maximum tolerated dose of phosmet in tthis
    strain of rat. Hepatocytes were isolated and prepared at 2 and 16 h;
    two independent experiments were carried out at each time. Groups of
    two animals received the vehicle or dimethylhydrazine dihydrochloride
    as a positive control. The mean net nuclear grain counts and the
    percentages of cells in repair were recorded. Phosmet did not induce
    DNA repair at either dose or time, whereas the positive control
    produced marked unscheduled DNA synthesis in comparison with vehicle
    controls (Mackay, 1996). 

    Comments

         In the study of delayed polyneuropathy, a dose of 600 mg/kg bw
    was given to 24 hens, this dose being greater than the experimentally
    determined LD50. There was no evidence that phosmet could produce
    clinical signs of delayed polyneuropathy or significantly inhibit
    neuropathy target esterase.

         The ability of phosmet (96.4% pure) to induce unscheduled DNA
    synthesis in the liver of male rats  in vivo was determined with
    doses of 32 or 50 mg/kg bw, the higher dose being the maximum
    tolerated dose of phosmet. Unscheduled DNA synthesis was not observed.
    The Meeting noted that the study on DNA binding had not been provided,
    but it concluded that no further characterization of mutagenicity was
    required. The Meeting considered that a further study in dogs would be
    unlikely to affect the overall evaluation.

         The ADI of 0-0.01 mg/kg bw allocated by the 1994 JMPR, which was
    based on a NOAEL of 1.3 mg/kg bw per day in a multigeneration study in
    rats and a safety factor of 100, was confirmed. 

         An acute RfD of 0.02 mg/kg bw was allocated on the basis of a
    NOAEL of 2 mg/kg bw per day in a study of developmental toxicity in
    rabbits (fetotoxicity) and a safety factor of 100. 

    Toxicological evaluation

     Levels that have no toxic effect (from 1994 monograph)

         Mouse:         25 ppm, equivalent to 4 mg/kg bw per day (two-year
                        study of carcinogenicity)

         Rat:           40 ppm, equal to 1.8 mg/kg bw per day (two-year
                        study of toxicity and carcinogenicity)
         Rat:           20 ppm, equal to 1.3 mg/kg bw per day
                        (two-generation study of reproductive toxicity)
                        5 mg/kg bw per day (maternal toxicity in study of
                        developmental toxicity)
                        15 mg/kg bw per day (study of developmental
                        toxicity)

         Rabbit:        5 mg/kg bw per day (maternal toxicity in study of
                        developmental toxicity)
                        2 mg/kg bw per day (study of developmental
                        toxicity)

     Estimate of acceptable daily intake for humans

              0-0.01 mg/kg bw

    Estimate of acute reference dose

              0.02 mg/kg bw

     Studies that would be useful for continued evaluation of the 
     compound

              Further observations in humans

    References

    Johnson, A.J. (1997) Phosmet acute delayed neurotoxicity study in the
    domestic hen. Unpublished report No. CTL/C/3123, dated 23 January
    1997, from Huntingdon Life Sciences, Huntingdon, Cambridgeshire,
    United Kingdom. Submitted to WHO by Gowan Co., Yuma, Arizona, USA.

    Mackay, J.M. (1996) Phosmet:  in vivo rat liver unscheduled DNA
    synthesis assay. Unpublished report No. CTL/P/5090, dated 23 September
    1996, from Central Toxicology Laboratory, Alderley Park, Cheshire,
    United Kingdom. Submitted to WHO by Gowan Co., Yuma, Arizona, USA.

    WHO (1996)  The WHO Recommended Classification of Pesticides by 
     Hazard and Guidelines to Classification 1996-1997 (WHO/PCS/96.3),
    International Programme on Chemical Safety, Geneva.
    


    See Also:
       Toxicological Abbreviations