Triphenyl phosphate

Specific investigations: other studies

Description of key information

Neurotoxicity is a potential adverse effect of many organosphosphates. In available studies in hens and cats pure triphenyl phosphate did neither induce immediate nor delayed neuropathy. The findings of a decreased activity of choline esterase and paralysis predominantly in cats and in older studies indicating neurotoxic potential were not reproduced in later studies and may be due to contamination of the tested samples by other organophosphorous esters. At the high doses of TPP used even small concentrations of impurities may have had sufficient activity.  Cytotoxicity investigations revealed no effects.

Additional information

Neurotoxicity is a potential adverse effect of many organophosphates. Some organo-phosphates also induce delayed neurotoxicity. Therefore TPP was tested for neurotoxicity in vivo and in vitro. It is recognised that the rat is a poor model for such delayed effects compared to the hen. Therefore the hen is according to OECD guideline 418 (acute exposure) the standard test model to detect these effects.

There are several reports of the investigation of TPP for such effects. The endpoints determined were the inhibition of choline esterases, neuropathy target esterase (NTE) and clinical observations in hens and cats and a functional observational battery in rats during subchronic exposure.

The earliest results regarding TPP were these being reported by Smith et al, 1932, who treated 4 hens orally with doses of 500 to 2000 mg/kg bw without any effects.

These findings were confirmed by Hine et al, 1956, at an oral dose of 1000 mg/kg bw. Animals were observed for their ability to walk for 14 to 36 days. Choline esterase was determined in plasma, brain, and spinal cord. Microscopic sections were examined from brain, spinal cord and sciatic nerve. The birds did not show signs of paralysis and no histological changes were detected. Choline esterase activity was reduced to 39 to 65% in plasma depending on the substrate.

In 1958, Henschler and Bayer described an experiment in hens using oral doses of up to 10000 mg/kg bw TPP (delivered in 2 -3days) in olive oil without any effect during the observation period.

Other studies (Aldridge et al, 1961) also found no sign of neurotoxicity after an oral dose of 500 mg/kg bw in hens after 3 weeks of observation, although they found a reduction of choline sterase in blood of 60%.

The low acute oral toxicity of pure TPP in hens is confirmed by later studies. In 1977, Johannsen et al, treated 9 hens twice daily with 5000 mg/kg bw for 3 days and after an interval of 18 days for another 3 days (total 60 g/kg bw). Animals sacrificed on day 42. Neither behavioural nor histological changes of nerve tissue were detected in any of the animals.

The activity of choline esterase was determined in a number of studies. Hine et al, 1956 found a severe depression of plasma choline esterase (without signs of paralysis) in hens, which was confirmed by Sutton et al, (1960) in mice after oral, intraperitoneal or inhalative administration and by Aldridge and Barnes (1961) in hens.

Sutton also reported that choline esterase activity was reduced after doses of 10 to 500 mg/kg bw in mouse whole blood to 87.1 to 30.4% of control. No cholinergic or other symptoms were reported. The partial inhibition of choline esterase activity in the whole blood was interpreted as a sign of absorption of TPP although a causal correlation was not proven.

Sutton at et al, (1960) described further experiments in rats, mice and guinea pigs using single doses of 3000 mg/kg bw in male and female animals leading to no mortality nor any clinical signs of toxicity at all. Additionally, Sutton et al, (1960), found inhibitory effects in human blood in vitro. At a concentration of 6X10E-5 Mol/L effects were most pronounced in human erythrocytes, human plasma and mouse whole blood with residual activities of 21, 40 or 57% resp. (unspecified period of incubation). At a concentration of 6X10E-7 Mol/L no inhibition was recorded.

Neuropathy target esterase (NTE), an enzyme involved in the development of delayed neuropathy after organophosphate exposure in animals, was determined by Johnson (1975) in hen brain homogenate. Johnson found a structure activity relationship (SAR) in a large series of organophosphorous compounds that indicates no neurotoxic potential for TPP. This SAR was confirmed by Johannsen et al, (1977).

Padilla et al, (1987) also found that TPP did not inhibit NTE in vitro in the microsomal fraction of rat brain and spinal cord tissues at concentrations of 1 to 10uM after 20 minutes of incubation. At 100uM NTE was inhibited to about 60% while positive control showed complete inhibition below 10uM.

Most of these studies were performed in hens, as this is the species of choice for organosphosphate induced neuropathy or in cats, as this species seems to be exceptionally sensitive to TPP. The major fault of many of these studies is that there are no reports of the purity of the tested samples. As these studies were performed before 1970 one has to assume that in many cases the samples were contaminated by other phosphoric acid esters due to the synthetic processes used during those years (Bayer, 2002).

Only one study employed high purity TPP (Wills et al, 1979). Two cats each were injected once subcutaneously with doses of 400, 700 mg/kg bw. One cat received 1000 mg/kg bw and two additional cats were employed as controls. At 400 mg/kg bw one cat stayed without findings while the other lost weight (31%) before recovering within 3 months. No signs of unusual weakness or ataxia were seen. At 700 mg/kg bw both animals became anorexic with watery faeces and prostration within several days. Histology of various organs revealed generalised vascular damage with perivascular edema in many tissues. The epithelial lining of the colon was lost. The livers showed fatty change and sinusoidal dilation. No changes were seen in the kidneys. Sections from 11 levels of the nervous system from cortex to peripheral nerve were examined and did not provide any evidence of axon degeneration, demyelination or any other pathological change. The cat given 1000 mg/kg bw became anorexic within one week, was unstable to rise from the floor 3 weeks after treatment, when it was killed. It had lost 48% of its original weight. Sections of the nervous tissue did not show any evidence of neuronal damage.

In a 4 -month study in rats, Sobotka et al (1986), determined the influence of dietary treatment with TPP at levels of 0, 0.25, 0.5, 0.75 and 1% corresponding to 161 to 711 mg/kg bw on the nervous system of male rats (10/group). In addition to standard clinical observations the neurotoxicity was assessed in open field, accelerating rotarod, forelimb grip strength and negative geotaxis examinations. These parameters were determined 4 times at the end of each month of treatment. Additionally body weights and food consumption were recorded weekly. No adverse effects were noted in any of the neurotoxicity parameters. Body weights were dose dependently reduced at 0.5% and 1% TPP.

Saboori et al, (1991) and Mandel et al, (1989) reported that TPP efficiently inhibits the non-specific esterase activity of human monocytes in vitro. This effect was discovered incidentally during haematological investigations in TPP exposed workers and not associated with any detectable health effect. There was no effect of TPP on the number and function of monocytes in a group of 38 workers similarly exposed to TPP and investigated by Emmett at al, (1985). The acetyl choline esterase of erythrocytes was depressed only marginally in these workers.