Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea

Endpoint summary

Description of key information

THPC-urea, like THPC is characterized by primary local toxicity (irritation or corrosion) depending on the concentration applied. The route of exposure is accountable for substance related effects. General systemic effects are considered to be secondary and maybe caused by repeated local adverse effects.
Ingestion of THPC in subchronic or chronic studies caused hepatocytic vacuolar degeneration in rats and mice and decreased survival in rats. 
The liver was the organ affected by oral exposure. Subchronic studies: NOEL = 3.75 mg/kg-d; Chronic studies LOEL = 3.75 mg/kg-d.
In pregnant rabbits, the subacute oral local NOAEL = 2 mg/kg bw /day (low dose) based on changes of gastrointestinal mucosal lining whereas the systemic NOAEL = 30 mg/kg bw/day (mid dose) based on body weight loss. (Factor 15)
Repeated dermal exposure studies revealed primary local toxicity of THPC. 
- In mice, the chronic dermal local NOEL = 0.5 mg/cm2 in mice (corresponding to 100 mg/kg bw/day) based on minimal dermal irritation (no systemic effects).
- in rats, the subacute dermal local LOAEL = 15 mg/cm2 based on severe skin necrosis (corresponding to a systemic LOAEL = 450 mg/kg bw/day based on body weight loss.
- in rabbits, the subacute dermal local LOAEL = 9.4 mg/cm2 severe skin necrosis (corresponding to a systemic LOAEL = 50 mg/kg bw/day based on declined condition.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

repeated dose toxicity: oral

Remarks:

other: subacute gavage in pregnant rabbits

Type of information:

experimental study

Adequacy of study:

key study

Study period:

190-1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Experimental data (GLP, per guideline)

Qualifier:

according to guideline

Guideline:

other: OECD 414 (Prenatal development toxicity study)

Deviations:

no

GLP compliance:

yes

Limit test:

no

Species:

rabbit

Strain:

New Zealand White

Sex:

female

Route of administration:

oral: gavage

Vehicle:

water

Remarks:

dosing volume: 5ml/kg bw

Details on oral exposure:

Each female was mated with 1 study male. After coitus females were injected with 10 I.U. chorionic gonadotrophin i.v. The day of insemination is taken as day 0 of gestation. Duration of treatment / exposure: day 7 to 19 of gestation = 13 consecutive days

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

day 7 to 19 of gestation = 13 consecutive days

Frequency of treatment:

once daily

Remarks:

Doses / Concentrations:
10 mg/kg bw/day
Basis:
nominal in water

Remarks:

Doses / Concentrations:
30 mg/kg bw/day
Basis:
nominal in water

Remarks:

Doses / Concentrations:
100 mg/kg bw/day
Basis:
nominal in water

No. of animals per sex per dose:

16

Control animals:

yes, concurrent vehicle

Details on study design:

for the dose range finding study see IUCLID chapter 7.8.2 Developmental Toxicity

Positive control:

No

Observations and examinations performed and frequency:

Morbidity/mortalitiy: twice daily; Clinical observations: once daily; Body weight: on day 0, 7, 8, 9, 12, 19, 24 and 29 of gestation; food intake: 0 to 3, 3 to 7, 7 to 8, 8 to 9, 9, to 12, 12 to 15, 15 to 17, 17 to 19, 19 to 21, 21 to 24, 24 to 27 and 27 to 29 of gestation.

Sacrifice and pathology:

3.8.1 Necropsy
Moribund females were killed by an intravenous injection of sodium pentobarbitone solution (200 mg/mL). Dead/killed females were necropsied in order to establish, where possible, the cause of death/moribund condition. Pregnancy status was recorded.
Surviving females were killed on day 29 after mating, in random group order, by an intravenous injection of sodium pentobarbitone solution (200 mg/mL) and examined macroscopically.
3.8.2 Uterine/implantation data
The ovaries and uteri were removed and examined and the following data recorded:
pregnancy status, number of corpora lutea, numbe~ and intrauterine position of implantations subdivided into:
live foetuses, early intrauterine deaths, late intrauterine deaths, dead foetuses
Early intrauterine deaths were classified as those which showed decidual or placental tissue only. Late intrauterine deaths showed embryonic or foetal tissue in addition to placental tissue. Oead foetuses were class1fied as those which appeared to have died shortly before necropsy.
Implantations were allocated numbers relating to their position in utero as follows:
Left horn: L1O to L1
Right horn: R10 to R1
The uteri of apparently non-pregnant animals were immersed in a 10% ammonium sulphide solution to reveal any evidence of implantation.
3.8.3 Foetal data
Live foetuses were killed by an intracardiac injection of sodium pentobarbitone solution.
Foetuses were weighed individually and examined externally. Approximately one half of ttte foetuses in each litter (selected by systematic sampling/ were decapitated by a cut through the neck. The heads were placed in Bouin's fluid for at least 6 weeks to allow fixation and partial decalcification. They were then transferred to 70% industrial methylated spirit and sections of the head examined. The remaining portion of the decapitated foetuses
and all intact foetuses were dissected. sexed anc the viscera examined. Following examination the foetuses were eviscerated and the carcasses placed in 70% industrial methylated spirit for approximately 24 hours. After fixation, the heads of the intact foetuses were sliced through the line of the frontoparietal suture and the brain was examined for visible abnormalities.
Following examination. the carcasses were processed to stain the ossified skeleton (Alizarin technique). the skeleton was examined then preserved and stored in absolute glycerol (containing thymol crystals to prevent fungal growtn). The head sections of the decapitated foetuses were preserved in
Bouin's fluid and stored in plast;c vials.
Foetal abnormalities were recorded as mal formations (rare and/or potentially lethal) and variations (commonly occurring non-iethal abnormalities).

Statistics:

3.9 Oata evaluation
3.9.1 Data processing
Data were processed. where appropriate. to give litter mean values. group mean values and standard deviations.
Some tables and appendices presented in thc report are computer generated. The group mean and individual data are each generated from the values held on a data base and rounded appropriately for inclusion in the report. As a consequence calculation of group mean data from the individual data presented in the report may, in same instances. yield a minor variation in the last decimal place/s.
3.9.2 Calculations
Group mean calculations in some instances (foetal weight) were based on litter mean rather than individual foetal data.
Group me an percentages were calculated us;ng group totals.
Percentage pre-implantation lass was calculated as:
number of corpora lutea - number of implantations
-------------------x 100
number of corpora lutea

Percentage post-implantation loss was calculated as:
number of implantations - number of live foetuses
-------------------- x 100
number of implantations

Percentage male foetuses was calculated as;
number of male foetuses
-------------- x 100
number of foetuses of detenmined sex

3.9.3 Statistical evaluation
The following parameters were analysed using the procedures stated in Appendix 2 of the protocol (Appendix 10):
body weight and body weight gain, food intake, corpora lutea, implantations, foetuses, pre-implantation loss, post-implantation loss, litter weight,foetal weight, percentage male pups, malformations, variations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Coloured urine was observed oceasionally in 11/16 animals in the high dose group. but was not apparent at the lower dose levels investigated. Aslightly higher incidence of rhinorrho.ea was observed in the intermediate dose group but was considered unlikely to have been an effeet of treatment as there was no effeet at the higher dose level. Other commonly occuring clinical signs were observed in all groups but their nature and incidence did not suggest an effrct of treatment. There were no unusual lesions at necropsy and no effect of treatment on their intergroup distribution. One female in the intermediate dose group was found dead on day 20 of gestation having shown abnormal respiration. Necropsy revealed lesions indicative of an intubation error. Abnormal respiration was also observed in one animal from the high dose group which was killed on day 13 of gestation. Dark eyes and coloured urine were also observed in this animal. At necropsy, mottling of the liver and darkening of the lungs were observed and all implantations were non-viable. An effect of tl"eatment was considered unlikely, however, as these signs were not observed in other treated animals.

Mortality:

mortality observed, treatment-related

Description (incidence):

Coloured urine was observed oceasionally in 11/16 animals in the high dose group. but was not apparent at the lower dose levels investigated. Aslightly higher incidence of rhinorrho.ea was observed in the intermediate dose group but was considered unlikely to have been an effeet of treatment as there was no effeet at the higher dose level. Other commonly occuring clinical signs were observed in all groups but their nature and incidence did not suggest an effrct of treatment. There were no unusual lesions at necropsy and no effect of treatment on their intergroup distribution. One female in the intermediate dose group was found dead on day 20 of gestation having shown abnormal respiration. Necropsy revealed lesions indicative of an intubation error. Abnormal respiration was also observed in one animal from the high dose group which was killed on day 13 of gestation. Dark eyes and coloured urine were also observed in this animal. At necropsy, mottling of the liver and darkening of the lungs were observed and all implantations were non-viable. An effect of tl"eatment was considered unlikely, however, as these signs were not observed in other treated animals.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

In the high dose group, 13/16 females lost weight between day 7 and day 9 of gestation. The reduction in group mean weight.was statistically significant (p<.05. Kruskal-Wallis, Wilcoxon rank sum test). A further reduction in weight gain was also observed

Food efficiency:

effects observed, treatment-related

Description (incidence and severity):

Food intake of the high dose group was significantly lower than controls over the treatment period (p<.05,days 7 to 9, p<.Ol days 9 to 12, p<.OOl days 12 to 19. Analysis of variance, Dunnett1s test), but comparable thereafter.

Details on results:

Body weight
In the high dose group, 13/16 females lost weight between day 7 and
day 9 of gestation. The reduction in group mean weight.was
statistically significant (p<.05. Kruskal-Wallis, Wilcoxon rank sum
test). A further reduction in weight gain was also observed in the
latter half of the dosing period (days 12 to 19, p<.Ol, Analysis of
variance, Dunnettls test). After treatment ceased, we;ght gain was
comparable to controls.
We;ght gain of the low and intermediate dose groups was comparable
to controls over the treatment period. Subsequently, weight gain
of these groups was marginally lower than controls but the
reduction was not statistically significant and an association with
treatment was considered unlikely.
Food intake
Food intake of the high dose group was significantly lower than
controls over the treatment period (p<.05,days 7 to 9, p<.Ol days 9
to 12, p<.OOl days 12 to 19. Analysis of variance, Dunnett1s test),
but comparable thereafter.
Food intake of the low and intermediate dose groups was comparable
to controls throughout gestation.
4.6 Caesarian

Dose descriptor:

NOAEL

Remarks:

maternal

Effect level:

30 mg/kg bw/day (nominal)

Based on:

test mat.

Remarks:

(65-68%)

Sex:

female

Basis for effect level:

other: body weight loss, decreased body weight gain and decreased food consumption at 100 mg/kg/d

Critical effects observed:

not specified

Caesarian data

Uterine/implantation data

Pregnancy rate was 93.8 or 100% in all groups. One animal in

the low dose group showed total resorption but no effect of

treatment was indicated as one incidence of total resorption

was also observed in the control group.

The number of corpora lutea per doe was generally comparable

amongst all groups. The number of implantations was lower

than controls in the high dose group, giv;ng rise to a

signif1cant dose response (p<.05 Terpstra-Jonckheere test) in

pre-implantation 1055. It is possible that there may have

been a relationship between the increased pre-implantation

1055 observed in the high dose group and the weight lasses

observed in this group at the start of treatment but as

implantation 1s expected to occur before day 7 of gestation

no conclusive association can be made. In addition. there

was no direct relationship between pre-imp1antation lass and

weight loss in individual animals.

Values for the 10w and intermediate dose groups, a1though

marginal1y higher than contro1s. were within the expected

range (7.8 to 14.6), and no effect of treatment was

concluded.

Treatment did not adversely affect post-implantation loss.

No dosage-related trends were apparent and all values were

within the expected range (1.7 to 11.9). Litter size was

comparable to the normal range in all groups (7.4 to 8.2).

4.6.2 Foetal data (Table 5.2. Appendices 5.2 and 9.1.1)

Percentage male foetuses pEr litter (sex ratio) was

comparable to controls in the high dose group. The unusua11y

high value for the low dose group was not statistically

significant and was considered to be of little toxicological

significance given the normal value for the intermediate dose

group.

litter weight generally reflected litter size a~d no effect

of treatment was concluded. There was no adverse effect of

treatment on foetal weight: values for all treated groups

were comparable to cr exceeded controls.

Foetal defect data

There was no effect of treatment on the number of foetuses

showing external/visceral and/or skeletal mal formations. The

incidence in all treated groups was lower than controls and

there were no trends in the type of mal formations observed.

The incidence of foetuses showing external/visceral

variations was comparable to controls in the low dose group,

marginally higher in the intermediate dose group and slightly

higher in the high dose group. This trend was statistically

significant (p<.05, Terpstra-Jonckheere test) but its

toxicological significance is questionable fiS the variations

involved (abnormal common carotid and agenesis of the

intermediate lung lobe) are known to occur spontaneously in

this strain of rabbit (Appendix 9.2). In the absence of any

correlation between the occurrence of these variations within

a litter and maternal weight loss. and in the absence of

other thoracic mal formations they are considered unlikely to

be indicative of a teratogenic even~.

There was no effect of treatment on the overall incidence of

skeletal variations. There were no dosage-related trends in

the incidence of any specific skeletal variations but the

proportion of foetuses with extra thoracic ribs was higher in

all treated groups than controls. An association with

treatment is inconclusive. however. as this variation has

been observed at a similar incidence (68.7%) in the control

group of a subsequent study.

Conclusions:

Treatment with the technical substance at a nominal dose level of 100 mg/kg/day elicited minimal maternal toxicity, characterised by colouration of the urine, reduced weight gain and reduced food intake.
There were, however, no adverse foetal effects which could conclusively be related to treatment.
There was no effect of treatment, maternal or foetal, at nominal dose levels of 10 or 30 mg/kg/day.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

30 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Quality of whole database:

Sufficient (AF=1) because underestimation of the risk is very unlikely:
- data obtained with Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea, CAS 27104-30-9 be neans of OECD 414 study:
13days oral exposure of pregnant rabbits revealed an systemic NOAEL = 30 mg/kg bw/day based on body weight loss. The local NOAEL = 2 mg/kg bw /day based on local changes of gastrointestinal mucosal lining)
- supporting read-across from Tetrakis(hydroxymethyl)phosphonium chloride (THPC, CAS 124-64-1), which has a higher toxicological potential: NTP studies are fully reliable data, though in rat and mouse. Unfortunately local effects were ignored in those oral studies. Subchronic/chronic oral toxicity data in non-rodents are not available and considered to be not relevant because of exposure considerations.

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Link to relevant study records

Reference

Endpoint:

chronic toxicity: dermal

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1978

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The read-across approach is based on the assumption, that Tetrakis(hydroxymethyl)phosphonium chloride (THPC, CAS 124-64-1), as part of the technical product 'Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea' (THPC-urea, CAS 27104-30-9), is the relevant, most hazardous compound. The (local) mode of action based on direct chemical reactivity is the same in both compounds. Additionally the read-across approach is based on the worst-case assumption that the reaction mixture THPC-urea is as reactive as THPC. In the confirmatory information it becomes obvious, that THPC is at least 6 times more hazardous than THPC, oligomeric reaction products with urea. For the detailed justification of the read-across hypothesis see “overall remarks, attachments”. When considering both the read-across hypotheses, the direct read-across from data obtained with technical THPC (80%) onto technical THPC-urea (65-68%) do not lead to an underestimation of the hazard. No further uncertainty factor, e.g. AF=2 for read-across, is considered to be necessary for deriving a DNEL(THPC-urea) from NOAEL(THPC). Published test report from Cancer Research and US-EPA assessment report thereof. Evaluated data from a reliable secondary source (US-CPSC).

Qualifier:

no guideline followed

Guideline:

other: one treatment group of 60 female mice (limit test at MTD, dermal) four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

Deviations:

no

Principles of method if other than guideline:

Limit test at MTD: one treatment group of 60 female mice; skin application 3 times weekly for 496 days (mouse chronic)
four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

GLP compliance:

not specified

Limit test:

yes

Species:

mouse

Strain:

ICR

Sex:

female

Details on test animals or test system and environmental conditions:

random-bred female ICR/Ha Swiss mice (ARS/Sprague-Dawley Division, The Mogul Corp., Madison, Wis.) were acclimated to laboratory conditions
for 2 weeks, and treatments were begun when they were 6 to 8 weeks old. The mice were housed 6/cage on sterile hardwood chips (Betta Chip; Fisher & Son, Bound Brook, N. J.) in stainless steel cages, fed Purina laboratory chow and water ad libitum, and weighed monthly. The animal rooms were maintained at 22 to 24°C.

Type of coverage:

not specified

Vehicle:

other: aceton-water (9:1)

Details on exposure:

external dose: 2 mg THPC per 0.2 ml aceton-water (9:1)
The dosages used in all experiments were based on short-term toxicity evaluations (4 to 6 weeks); the highest possible doses that gave
minimal cytotoxic effects, as determined by histopathological examination, were used for the chronic exposures.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

skin application 3 times weekly for 496 days

Frequency of treatment:

skin application 3 times weekly for 496 days

Remarks:

Doses / Concentrations:
2 mg/application
Basis:
nominal per unit area

No. of animals per sex per dose:

one treatment group of 60 female mice (limit test at MTD, dermal)
four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

Control animals:

yes, concurrent no treatment

yes, concurrent vehicle

Positive control:

No addition of tumor inductor or promotor.
Study hypothesis is, the possible liberation of formaldehyde and hydrochloric acid from free THPC in the fabric. Formaldehyde and hydrochloric acid are known to be in equilibrium with the animal and human carcinogen bis(chloromethyl)ether (Van Duuren 1978).

Observations and examinations performed and frequency:

All compounds were used in well-ventilated hoods, and the mice were housed there for at least 2 hr after treatment. All skin treatments were
continued for the duration of the experiment. Animals were examined regularly and scored, and the findings were charted once monthly.

Sacrifice and pathology:

Animals in poor health or with large tumor masses were killed. Except for the cranial region, animals were completely autopsied at the end of the
experiment or at death. Samples of all abnormal-appearing tissues and organs were excised for histopathological diagnosis. In addition, routine
histopathology was performed on skin, liver, and kidney in all animals dying during the test period and in 20% of all animals killed at the end of the
test. All tissue sections were fixed in 10% formalin, proc essed, blocked in paraffin, and stained with hematoxylin and eosin for histopathological
examination. Included in the protocols were groups given vehicle only; no treatment groups were included in the protocols.

Other examinations:

Skin lesions were diagnosed clinically as papillomas when they reached 1 cu mm in size and persisted for 30 days or more or when they were confirmed histopathologically.

Statistics:

The p values given in the table were assigned by means of computation of x2 analyses in which the tumor
incidences in the test groups were compared with a composite of 4 no-treatment control groups that were used for
these tests as well as for other concurrent experiments (249 mice).

Clinical signs:

effects observed, treatment-related

Dermal irritation:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

THPC was inactive in induction of skin tumors in skin painting studies.
THPC is a quaternary compound. Such compounds do not cross membranes very readily, therefore, they are not absorbed via the skin and poorly
absorbed from mucuous membranes. The chloride salts of quaternary bases are not likely to generate active chloride ions that could then help to
form bischloromethyl ether from the THPC.

Dose descriptor:

other: maximal tolerated dose (dermal)

Effect level:

2 other: mg/0.2 mL external dose

Based on:

act. ingr.

Sex:

female

Basis for effect level:

other: minimal cytotoxic (local irritation) effect to mouse skin

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (actual dose received)

Based on:

act. ingr.

Sex:

female

Basis for effect level:

other: minimal effect on mouse skin, no systemic toxicity

Critical effects observed:

not specified

No significant increases in tumors were observed when THPC (2 mg/animal) in acetone:water was tested for carcinogenic activity

by dermal application 3 times/week for 496 days (VanDuuren et al. 1978). Of the 59 treated animals necropsied, 17 lung (papillary)

tumors and 1 forestomach (papilloma) were observed, compared to 90 lung (papillary) tumors and 7 forestomach tumors out of the

249 untreated mice necropsied (p>0.05). Of the 29 mice treated with acetone:water (90:10) and examined, there were 7 lung

(papillary) tumors.

THPC was inactive in induction of skin tumors in skin painting studies.

The maximal tolerated dose (MTD dermal) of technical THPC is 2 mg/mouse per day. (Mouse bw 18g, body surface 75cm2)

external dose= 2mg/0.2ml (acteon:water 9:1)=10g/L (1%)

MTD=NOAEL (dermal, local) = 2mg/10% body surface mouse= 2mg/ 7.5 cm2 = 0.27 mg/cm2.

NOAEL (dermal, systemic)= 2mg/18g mouse= 100mg/kg bw/day

Conclusions:

The dermal toxicity study did not provide indications for systemic toxicity. Only slightly local effects were observed at the dose level applied.
In this chronic dermal test, based on minimal local irritation, a MTD of 2 mg THPC/animal=2mgTHPC/7.5cm2 = 0.25 mg/cm2, corresponding to 100 mg/kg bw/day (vehicle: aceton:water=9:1) what was identified to be the NOAEL in mice.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

100 mg/kg bw/day

Study duration:

chronic

Species:

mouse

Repeated dose toxicity: dermal - local effects

Link to relevant study records

Reference

Endpoint:

chronic toxicity: dermal

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

1978

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The read-across approach is based on the assumption, that Tetrakis(hydroxymethyl)phosphonium chloride (THPC, CAS 124-64-1), as part of the technical product 'Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea' (THPC-urea, CAS 27104-30-9), is the relevant, most hazardous compound. The (local) mode of action based on direct chemical reactivity is the same in both compounds. Additionally the read-across approach is based on the worst-case assumption that the reaction mixture THPC-urea is as reactive as THPC. In the confirmatory information it becomes obvious, that THPC is at least 6 times more hazardous than THPC, oligomeric reaction products with urea. For the detailed justification of the read-across hypothesis see “overall remarks, attachments”. When considering both the read-across hypotheses, the direct read-across from data obtained with technical THPC (80%) onto technical THPC-urea (65-68%) do not lead to an underestimation of the hazard. No further uncertainty factor, e.g. AF=2 for read-across, is considered to be necessary for deriving a DNEL(THPC-urea) from NOAEL(THPC). Published test report from Cancer Research and US-EPA assessment report thereof. Evaluated data from a reliable secondary source (US-CPSC).

Qualifier:

no guideline followed

Guideline:

other: one treatment group of 60 female mice (limit test at MTD, dermal) four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

Deviations:

no

Principles of method if other than guideline:

Limit test at MTD: one treatment group of 60 female mice; skin application 3 times weekly for 496 days (mouse chronic)
four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

GLP compliance:

not specified

Limit test:

yes

Species:

mouse

Strain:

ICR

Sex:

female

Details on test animals or test system and environmental conditions:

random-bred female ICR/Ha Swiss mice (ARS/Sprague-Dawley Division, The Mogul Corp., Madison, Wis.) were acclimated to laboratory conditions
for 2 weeks, and treatments were begun when they were 6 to 8 weeks old. The mice were housed 6/cage on sterile hardwood chips (Betta Chip; Fisher & Son, Bound Brook, N. J.) in stainless steel cages, fed Purina laboratory chow and water ad libitum, and weighed monthly. The animal rooms were maintained at 22 to 24°C.

Type of coverage:

not specified

Vehicle:

other: aceton-water (9:1)

Details on exposure:

external dose: 2 mg THPC per 0.2 ml aceton-water (9:1)
The dosages used in all experiments were based on short-term toxicity evaluations (4 to 6 weeks); the highest possible doses that gave
minimal cytotoxic effects, as determined by histopathological examination, were used for the chronic exposures.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

skin application 3 times weekly for 496 days

Frequency of treatment:

skin application 3 times weekly for 496 days

Remarks:

Doses / Concentrations:
2 mg/application
Basis:
nominal per unit area

No. of animals per sex per dose:

one treatment group of 60 female mice (limit test at MTD, dermal)
four no-treatment control groups that were used for these tests as well as for other concurrent experiments (249 mice).

Control animals:

yes, concurrent no treatment

yes, concurrent vehicle

Positive control:

No addition of tumor inductor or promotor.
Study hypothesis is, the possible liberation of formaldehyde and hydrochloric acid from free THPC in the fabric. Formaldehyde and hydrochloric acid are known to be in equilibrium with the animal and human carcinogen bis(chloromethyl)ether (Van Duuren 1978).

Observations and examinations performed and frequency:

All compounds were used in well-ventilated hoods, and the mice were housed there for at least 2 hr after treatment. All skin treatments were
continued for the duration of the experiment. Animals were examined regularly and scored, and the findings were charted once monthly.

Sacrifice and pathology:

Animals in poor health or with large tumor masses were killed. Except for the cranial region, animals were completely autopsied at the end of the
experiment or at death. Samples of all abnormal-appearing tissues and organs were excised for histopathological diagnosis. In addition, routine
histopathology was performed on skin, liver, and kidney in all animals dying during the test period and in 20% of all animals killed at the end of the
test. All tissue sections were fixed in 10% formalin, proc essed, blocked in paraffin, and stained with hematoxylin and eosin for histopathological
examination. Included in the protocols were groups given vehicle only; no treatment groups were included in the protocols.

Other examinations:

Skin lesions were diagnosed clinically as papillomas when they reached 1 cu mm in size and persisted for 30 days or more or when they were confirmed histopathologically.

Statistics:

The p values given in the table were assigned by means of computation of x2 analyses in which the tumor
incidences in the test groups were compared with a composite of 4 no-treatment control groups that were used for
these tests as well as for other concurrent experiments (249 mice).

Clinical signs:

effects observed, treatment-related

Dermal irritation:

effects observed, treatment-related

Mortality:

mortality observed, treatment-related

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Histopathological findings: neoplastic:

effects observed, treatment-related

Details on results:

THPC was inactive in induction of skin tumors in skin painting studies.
THPC is a quaternary compound. Such compounds do not cross membranes very readily, therefore, they are not absorbed via the skin and poorly
absorbed from mucuous membranes. The chloride salts of quaternary bases are not likely to generate active chloride ions that could then help to
form bischloromethyl ether from the THPC.

Dose descriptor:

other: maximal tolerated dose (dermal)

Effect level:

2 other: mg/0.2 mL external dose

Based on:

act. ingr.

Sex:

female

Basis for effect level:

other: minimal cytotoxic (local irritation) effect to mouse skin

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (actual dose received)

Based on:

act. ingr.

Sex:

female

Basis for effect level:

other: minimal effect on mouse skin, no systemic toxicity

Critical effects observed:

not specified

No significant increases in tumors were observed when THPC (2 mg/animal) in acetone:water was tested for carcinogenic activity

by dermal application 3 times/week for 496 days (VanDuuren et al. 1978). Of the 59 treated animals necropsied, 17 lung (papillary)

tumors and 1 forestomach (papilloma) were observed, compared to 90 lung (papillary) tumors and 7 forestomach tumors out of the

249 untreated mice necropsied (p>0.05). Of the 29 mice treated with acetone:water (90:10) and examined, there were 7 lung

(papillary) tumors.

THPC was inactive in induction of skin tumors in skin painting studies.

The maximal tolerated dose (MTD dermal) of technical THPC is 2 mg/mouse per day. (Mouse bw 18g, body surface 75cm2)

external dose= 2mg/0.2ml (acteon:water 9:1)=10g/L (1%)

MTD=NOAEL (dermal, local) = 2mg/10% body surface mouse= 2mg/ 7.5 cm2 = 0.27 mg/cm2.

NOAEL (dermal, systemic)= 2mg/18g mouse= 100mg/kg bw/day

Conclusions:

The dermal toxicity study did not provide indications for systemic toxicity. Only slightly local effects were observed at the dose level applied.
In this chronic dermal test, based on minimal local irritation, a MTD of 2 mg THPC/animal=2mgTHPC/7.5cm2 = 0.25 mg/cm2, corresponding to 100 mg/kg bw/day (vehicle: aceton:water=9:1) what was identified to be the NOAEL in mice.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

0.25 mg/cm²

Study duration:

chronic

Species:

mouse

Quality of whole database:

Sufficient (AF=1) because underestimation of the risk is very unlikely:
- no data of the substance (Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea, CAS 27104-30-9) are available.
- instead read-across from Tetrakis(hydroxymethyl)phosphonium chloride (THPC, CAS 124-64-1), which has a higher toxicological potential
- The MTD from a chonic test with THPC in mice is reliable data. No dermal local NOAEL for other species is known (neither THPC-urea nor THPC). For the endpoint local irritation no further hazard assessment is considered to be necessary and the data basis quality is sufficient (AF=1).

Additional information

No RDT studies with Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea, CAS 27104-30-9 are available, with the exception of a teratology study in pregnant rabbits.

The read-across of data from RD studies with THPC is based on the assumption, that Tetrakis(hydroxymethyl)phosphonium chloride (THPC) as component of the technical substance 'Tetrakis(hydroxymethyl)phosphonium chloride, oligomeric reaction products with urea' (THPC-urea) is the relevant, compound of concern. Additionally the read-across approach is based on the worst case assumption that THPC-urea (68%) is as reactive as THPC (80%). (Frankly speaking THPC is more hazardous than THPC-urea, but this cannot be quantified)

No underestimation of hazard/risk is possible when a direct read-across of data obtained with technical THPC (80%) onto technical THPC-urea (65-68%) is performed. No further uncertainty factor, e.g. AF=2 for read-across, is considered to be necessary for deriving a DNEL(THPC-urea) from an NOAEL(THPC).

THPC-urea, like THPC is characterized by primary local toxicity (irritation or corrosion) depending on the concentration applied. The route of exposure is accountable for substance related effects. General systemic effects are considered to be caused by severe and/or repeated local adverse effects.

ACGIH, 2005: Although THPC caused local skin irritation in mice, rats and rabbits, there is no evidence to suggest a systemic effect.Hence, a skin notation is not warranted.

Justification for selection of repeated dose toxicity inhalation - systemic effects endpoint:
exposure based waiving

Justification for selection of repeated dose toxicity inhalation - local effects endpoint:
exposure based waiving

Justification for selection of repeated dose toxicity dermal - systemic effects endpoint:
systemic toxicity is assessed to be secondary to local toxicity.

Justification for selection of repeated dose toxicity dermal - local effects endpoint:
local irritation is considered to be the primary toxic effect of the substance

Repeated dose toxicity: via oral route - systemic effects (target organ) digestive: liver

Repeated dose toxicity: dermal - systemic effects (target organ) other: skin

Justification for classification or non-classification

Classification for systemic effects, e.g. STOT RE cat 1 H372 (oral gavage, liver) is considered not necessary because of limited oral exposure under expected conditions of handling and use. Repeated oral exposure with an effective dose like in animal experiments is not a realistic exposure scenario for men (industrial professionals) because of the corrosive labelling of the substance. In an short-term gavage study with THPC-urea (see key study in IUCLID section 7.8.2), the effective level for systemic effects exceeds the no-effect level for local effects by factor 15.

Regarding the general public limited exposure (all routes) is claimed because of the multiple chemical modification during textile finishing.

Inhalation exposure is considered to be non-relevant because of expected conditions of industrial handling and use (tests waived).

In case of dermal exposure classification for acute local toxicity is necessary and considered to be sufficient: skin corrosion 1B, H314: Causes severe skin burns and eye damage.