1-[2-(2-chloroethoxy)phenylsulfonyl]-3-(4-methoxy-6-methyl-1,3,5-triazin-2-yl)urea;triasulfuron (ISO)

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 Oct 2011 to 01 Nov 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

Version / remarks:

2004

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Type of coverage:

open

Vehicle:

water

Duration of exposure:

24 hours

Doses:

- 33.3 % w/w aqueous slurry, nominally containing 66.7 g/kg
- 1/444 w/w dilutions, nominally containing 0.15 g/kg
- 1/888 w/w dilutions, nominally containing 0.075 g/kg

Time point:

24 h

Concentrate / Dilution:

concentrate

Dose:

33 % (w/w) , corresponding to 67.3 g substance/kg

Parameter:

percentage

Absorption:

0.04 %

Remarks on result:

other: dermal absorption = (stratum corneum (tape strips 3-5) + remaining epidermis + receptor fluid) + k * SD

Time point:

24 h

Concentrate / Dilution:

dilution

Dose:

1/444 (w/w), corresponding to 0.168 g substance/kg

Parameter:

percentage

Absorption:

1.8 %

Remarks on result:

other: dermal absorption = (stratum corneum (tape strips 3-5) + remaining epidermis + receptor fluid) + k * SD

Time point:

24 h

Concentrate / Dilution:

dilution

Dose:

1/888 (w/w), corresponding to 0.086 g substance/kg

Parameter:

percentage

Absorption:

4.4 %

Remarks on result:

other: dermal absorption = (stratum corneum (tape strips 3-5) + remaining epidermis + receptor fluid) + k * SD

STABILITY IN THE FORMULATIONS

(14C)-labelled test substance, when formulated as a 66.7 g/kg, 0.15 g/kg and 0.075 g/kg formulation, was shown to be stable by HPLC for a period of 1 day, which is a period of time longer than that used in the study.

Radiochemical purities of greater than 97 % were seen in all dose preparations both prior to application and one day post dose preparation.

 

PARTICLE SIZE

Particle size analysis of the dose preparations confirmed that the mean particle sizes were 6.95 μm (2.5 – 17.5 μm range), 7.43 μm (2.5 – 20 μm range) and 7.26 μm (2.5 – 21.25 μm range) for the 33.3 % w/w aqueous slurry and the 1/444 w/w and 1/888 w/w aqueous dilutions, respectively.

 

Table 1. Individual Distribution of the test substance from the 33.3 % w/w Aqueous Slurry of the Formulation Concentrate in the Test System

Test Compartment	Percentage of Dose Recovered (%)
	Cell 31	Cell 33	Cell 35	Cell 36	Cell 38	Cell 39
Donor chamber	0.004	0.002	0	0.011	0.02	0.02
Skin wash	108	99.9	114	110	113	109
Stratum corneum (Tape strips 1 & 2)	0.008	0.018	0.013	0.009	0.017	0.008
Stratum corneum (Tape strips 3 - 5)	0.004	0.007	0.005	0.006	0.01	NS
Remaining epidermis	0.007	0.02	0.009	0.01	0.007	0.044
Receptor fluid	0.012	0	0.007	0.001	0.004	0.002
Total recovered	108	100	114	110	113	109
Dermal absorption per cell	0.023	0.027		0.017		0.046

Stratum corneum = amount in tape strips; Remaining epidermis = epidermal tissue remaining after tape stripping

* (Cell 35 and 38 were excluded in the calculation of the mean absorption due to high recovery).

 

Table 2. Individual Distribution of the test substance from the 1/444 w/w Aqueous Slurry of the Formulation Concentrate in the Test System

Test Compartment	Percentage of Dose Recovered (%)
	Cell 1	Cell 2	Cell 3	Cell 6	Cell 7	Cell 15
Donor chamber	0	2.62	0	0.238	0	0
Skin wash	97.5	101	100	103	99.4	105
Stratum corneum (Tape strips 1 & 2)	0.183	0.231	0.423	0.287	0.315	0.064*
Stratum corneum (Tape strips 3 - 5)	0.183	0.358	0.367	0.127	0.166	0.025*
Remaining epidermis	1.57	1.44	0.973	0.804	0.553	0.284
Receptor fluid	1.35	0.149	0.075	0.139	0.094	0.056
Total recovered	101	106	102	104	100	105
Dermal absorption per cell	3.103	1.947	1.415	1.070	0.813	0.340

Stratum corneum = amount in tape strips; Remaining epidermis = epidermal tissue remaining after tape stripping

 

Table 3. Individual Distribution of the test substance from the 1/888 w/w Aqueous Slurry of the Formulation Concentrate in the Test System

Test Compartment	Percentage of Dose Recovered (%)
	Cell 5	Cell 9	Cell 10	Cell 19	Cell 26	Cell 28
Donor chamber	0.000*	0.817	0	0	0	0
Skin wash	106	102	93.9	93.4	103	99.9
Stratum corneum (Tape strips 1 & 2)	0.487	NS	0.989	0.534	0.172	0.381
Stratum corneum (Tape strips 3 - 5)	0.841	NS	0.985	0.414	0.210	0.457
Remaining epidermis	2.10	3.00	4.22	2.74	0.724	1.28
Receptor fluid	0.086	0.194	0.078	0.141	0.143	0.224
Total recovered	110	106	100	97.2	104	102
Dermal absorption per cell	3.027	3.194	5.283	3.295	1.077	1.961

Stratum corneum = amount in tape strips; Remaining epidermis = epidermal tissue remaining after tape stripping

Conclusions:

The penetration of the test substance from a 33.3 % w/w (corresponding to 67.3 g substance/kg) slurry, 1/444 w/w (corresponding to 0.168 g substance/kg) and 1/888 w/w (corresponding to 0.086 substance/kg) aqueous spray strength dilutions of the concentrate slurry, through human epidermis is very slow and the vast majority of the applied dose could be removed by gentle skin washing after 24 hours. The overall absorption of the test substance over 24 hours was 0.04, 1.8, and 4.4 %, for the 33.3 % w/w slurry, 1/444 w/w dilution, and 1/888 w/w dilution, respectively. These data predict that the systemic exposure to the test substance following dermal contact with this formulation would be minimal.

Executive summary:

The dermal absorption and distribution of the test substance from formulation concentrate was measured in human skin in a study conducted according to OECD TG 428 following GLP. Doses of (14C)-test substance were applied as a 33.3 % w/w aqueous slurry of the formulation concentrate (nominally, 66.7 g/kg) and as 1/444 w/w (nominally, 0.15 g/kg) and 1/888 w/w (nominally, 0.075 g/kg) aqueous spray strength aqueous dilutions of the slurry. The doses were applied at 10 μL/cm2 and left unoccluded for an exposure period of 24 hours. The absorption process was followed by taking samples of the receptor fluid (50 % v/v ethanol/water) at recorded intervals throughout the exposure period. The distribution of the test substance within the test system and a 24 hour absorption profile were determined, using liquid scintillation counting (LSC). The absorbed (systemically available) dose is calculated based on EFSA guidance on dermal absorption (2017). Based on the fact that the study has a duration of 24 hours, < 75 % of the permeation occurs in the first half of the study, and that the tape strips were analysed individually, the tape stips 1 + 2 were excluded from the absorption calculation. Dermal absorption = % in receptor fluid + % in skin sample. 

For the 33.3 % w/w aqueous slurry of the formulation concentrate slurry, the mean total recovery was 109 % of the dose applied. The overall dermal absorption of the test substance from the 33.3 % w/w aqueous slurry of the formulation concentrate through human epidermis was 0.04 % during the 24 hour exposure period. The amount absorbed at 24 hours was 0.028 μg/cm² (0.004 % of the dose applied). The amounts absorbed at 6, 8 and 12 hours were 0.021 μg/cm² (0.003 %), 0.021 μg/cm² (0.003 %) and 0.030 μg/cm² (0.004 %), respectively. These values were below the limit of quantitation (0.039 μg/cm², 0.006 %). The vast majority of the applied test substance (mean 109 %) was washed off the skin at 24 hours. 

For the 1/444 w/w aqueous dilution of the formulation concentrate slurry, the mean total recovery was 104 % of the dose applied. The overall dermal absorption of the test substance from the 1/444 w/w aqueous dilution of the formulation concentrate slurry through human epidermis was 1.8 % during the 24 hour exposure period. The amount absorbed at 24 hours was 0.0017 μg/cm² (0.103 % of the dose applied). The amounts absorbed at 6, 8 and 12 hours were 0.0005 μg/cm² (0.032 %), 0.0007 μg/cm² (0.040 %) and 0.0008 μg/cm² (0.050 %), respectively. The values for 6 and 8 hours were below the limit of quantitation (0.0008 μg/cm², 0.047 %).The vast majority of the applied test substance (mean 102 %) was washed off the skin at 24 hours. 

For the 1/888 w/w aqueous dilution of the formulation concentrate slurry, the mean total recovery was 103 % of the dose applied. The overall dermal absorption of the test substance from the 1/888 w/w aqueous dilution of the formulation concentrate slurry through human epidermis was 4.4 % during the 24 hour exposure period. The amount absorbed at 24 hours was 0.0012 μg/cm² (0.143 % of the dose applied). The amounts absorbed at 6, 8 and 12 hours were 0.0007 μg/cm² (0.080 %), 0.0007 μg/cm² (0.086 %) and 0.0009 μg/cm² (0.102 %), respectively. The values for 6 and 8 hours were below the limit of quantitation (0.0008 μg/cm², 0.090 %).The vast majority of the applied test substance (mean 99.8 %) was washed off the skin at 24 hours. 

The penetration of the test substance from a 33.3 % w/w slurry, 1/444 w/w and 1/888 w/w aqueous spray strength dilutions of the concentrate slurry, through human epidermis is very slow and the vast majority of the applied dose could be removed by gentle skin washing after 24 hours. The overall absorption of the test substance over 24 hours was 0.04, 1.8, and 4.4 %, for the 33.3 % w/w slurry, 1/444 w/w dilution, and 1/888 w/w dilution, respectively. These data predict that the systemic exposure to the test substance following dermal contact with this formulation would be minimal.

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

19 Oct 1986 to 20 May 1987

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Objective of study:

absorption

distribution

excretion

metabolism

Qualifier:

according to guideline

Guideline:

EPA OPP 85-1 (Metabolism and Pharmacokinetics)

Version / remarks:

1982

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Species:

rat

Strain:

Wistar

Remarks:

KFM-WIST (SPF)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 7 - 8 weeks of age
- Weight at study initiation: 150 - 200 g
- Housing: Groups of 3 rats in Makrolon type-4 cages with standard soft wood bedding during acclimatisation and treatment with unlabelled test substance. During the metabolism studies rats were held individually in all-glass metabolism cages.
- Diet: Ad libitum.
- Water: Tap water. Ad libitum
- Acclimatisation: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 37.5 - 70
- Air changes per hour: 10 - 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 19 Oct 1986 To: 20 May 1987

Route of administration:

other: Intravenous or oral

Details on exposure:

Male and female rats were orally or intravenously treated with a single high (300 mg/kg bw/day) or low (0.5 mg/kg bw/day) dose level of the test substance.

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

300 mg/kg bw (total dose)

Remarks:

Group 1: oral administration in males

Dose / conc.:

0.5 mg/kg bw (total dose)

Remarks:

Group 2: oral administration in males

Dose / conc.:

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

Remarks:

Group 3: oral administration in females

Dose / conc.:

300 mg/kg bw (total dose)

Remarks:

Group 4: oral administration in females

Dose / conc.:

0.5 mg/kg bw (total dose)

Remarks:

Group 5: intravenous administration in males

Dose / conc.:

0.5 mg/kg bw (total dose)

Remarks:

Group 6: intravenous administration in females

Dose / conc.:

0.5 mg/kg bw (total dose)

Remarks:

Group 7: oral administration in males

Dose / conc.:

0.5 mg/kg bw (total dose)

Remarks:

Group 8: oral administration in females

No. of animals per sex per dose / concentration:

Group 1: 5 males
Group 2: 5 males
Group 3: 5 females
Group 4: 10 females

Details on absorption:

The test substance was readily absorbed based on calculated excretion values (74.5 - 96.45 %).

Details on distribution in tissues:

The residual radioactivity levels found in organs/tissues, being after 96 hours below the limit of quantification (LOQ) for the low dose level (0.5 mg/kg) and in the order of LOQ (spleen, brain, fat, adrenal glands, thyroid gland) up to 0.88 mg/g (skin, chest) for the high dose level (300 mg/kg). After repeated administered at the low dose level, no radioactivity was retained in organs and tissues of male and female rats.

Details on excretion:

Excretion mainly via the urine representing in male rats, on average, 74.0 and 80.0 % of the radioactivity administered for the high and low dose level, respectively. The corresponding values in females were 74.4 and 85.1 %. Excretion via the faeces ranged for all experiments,  on average, from 0.3 to 13.9 %.

Metabolites identified:

yes

Details on metabolites:

Analysis of the metabolite patterns in urine, faeces and liver showed mainly the presence of the parent molecule, ranging from 65.7 - 84.9 % of the dose administered. Besides the test substance itself, 12 urinary metabolites (6.6 - 8.1 %), and 8 faecal meta­bolites (1.2 - 2.9 %) were identified.

BALANCE STUDY ON THE RAT AFTER SINGLE ORAL ADMINISTRATION

Balances and excretion patterns:

Excretion mainly proceeded via the urine representing in male rats, on average, 74.0 and 80.0 % of the radioactivity administered for the high and low dose level, respectively. The corresponding figures in females were 74.4 and 85.1 %.

The excretion patterns demonstrated a rapid elimination of absorbed radioactivity since already 24 hours after the administration, on average, 41.8 to 75 8 % of the radioactivity administered were found in the urine. Furthermore, it can be stated that urinary excretion was higher at the low dose levels than at the high dose levels. No influence of sex on absorption/urinary excretion was observed.

Excretion via the faeces ranged for all experiments,  on average, from 0.3 to 13.9 %. Additional radioactivity resulting from excretion via urine and faeces was found in the cage wash ranging, on  average, from 5.8 to 12.1 %.

Only very small amounts of radioactivity were found in the intestinal tracts (<0.1 %), in the residual carcass (<0.1-0.3 %), in organs/tissues (<0.1 %) and in the expired air (<0.01 %).

Total recoveries of radioactivity ranged, on average, from 92.9 to 101.8 % of the radioactivity administered.

In conclusion, the test substance was highly absorbed by male and female rats and thereafter rapidly and almost completely excreted.

Residual radioactivity in organs/ tissues/ blood:

Males high target dose level (300 mg/kg):

The residual radioactivity in organs/tissues, blood and plasma was low 96 hours after the administration of U-14C-labelled test substance. Except for liver (0.31 mcg/ g), kidney (0.68 mcg/g), pancreas (0.23 mcg/g), stomach (0.15 mcg/ g), intestinal tract (0.22 mcg/g), skin (0.29-0.88 mcg/g), carcass (0.41 mcg/g), blood (0.74 mcg/g) and plasma (1.09 mcg/g), all other organs and tissues were below or at the limit of quantification.

Females high target dose level (300 mg/kg):

Residual radioactivity in females was very similar to that observed in males, i.e. only liver, kidney, intestinal tract, ovary/uterus, skin, carcass, blood and plasma, significant but low amounts of radioactivity were detected (Tables 19 and 25). The reasons for these low amounts of residual radioactivity still found were mainly due to a not totally completed excretion process. Residual radioactivity observed in the skin was considered to be due to contaminations from urine and faeces.

Males and females: low target dose level (0.5 mg/kg):

The residual radioactivity levels in all organs/tissues, blood and plasma were below or at the limit of quantification. In conclusion, practically no radioactivity was retained 96 hours after the administration of the low dose level, and only small amounts were found at the high dose level in organs/tissues etc. involved in the excretion process. Therefore, it can be assumed that increasing the depuration period at the high dose level would also result in a complete elimination of residual radioactivity.

 

BALANCE STUDY ON THE RAT AFTER SINGLE INTRAVENOUS ADMINISTRATION

Balances and Excretion Patterns:

Urinary excretion was very rapid and high accounting already after 8 hours in males and females orally exposed to a target dose level of 0.5 mg/kg for 88.2 and 86.5 %, respectively. Finally after 96 hours, on average, 98.2 and 92.1 % were excreted via the urine by males and females, respectively.

Excretion via the faeces was in males and females 7.2 and 2.3 %, respectively. Additional radioactivity was collected in the cage washes (2.7 and 3.1 %).

Radioactivity found in the residual carcass and organs/tissues was 0.2 and <0.1 % of the radioactivity administered to male rats, respectively. For females, the corresponding figures were 0.1 and <0.1 %. Practically no radioactivity was found in the intestinal tract (<0.1 %) of males and females.

Total recoveries were, on average, 97.6 - 108.3 %.

When compared with the oral study, it can be stated that the total excretion and the ratio of  radioactivity excreted via the urine and faeces were of the same order of magnitude. Based on these findings, it can be concluded for the oral study that the amount of radioactivity appearing in the feces most probably originated from biliary excretion. Therefore, a complete absorption of the test substance from the gastro-intestinal tract can be assumed after oral administration.

Residual radioactivity in organs/ tissues/ blood:

When compared with the oral study, again no residual radioactivity was found in organs, tissues and in blood/plasma, i.e. all values were below the limit of quantification.

 

EXCRETION OF RADIOACTIVITY AFTER REPEATED (14X) ORAL ADMINISTRATION OF THE UNLABELLED TEST SUBSTANCE FOLLOWED BY A SINGLE ORAL ADMINISTRATION OF THE 14C­ LABELLED TEST SUBSTANCE

Excretion patterns:

Males: On average, 91.5 and 7.9 % of the radioactivity administered were excreted via the urine and faeces, respectively. Further, mainly urinary radioactivity was collected via the cage wash (4.4 %). Therefore, the total excreted radioactivity was 103.8 %. Only negligible amounts of radioactivity were found in the intestinal tract, residual carcass and organs/tissues. The total recovery was, on average, 104.0 %.

Females: Here, 82.0, 6.6 and 14.4 % of the radioactivity administered were collected in the urine, faeces and cage wash. Based on these figures, total excretion accounted for 103.0 %. Residual radioactivity in the intestinal tract, residual carcass and organs/tissues ranged from <0.1 - 0.1 %. The total recovery was, on average, 103.1 %.

When compared with the corresponding single oral administration study, a more rapid absorption/ elimination of radioactivity could be stated, i.e. absorption/elimination half-lives were about 8 hours and 24 hours for the repeated oral and single oral route of administration, respectively. These data indicate an adaptation of the rat to the test article after repeated oral administration resulting in a more pronounced urinary excretion rate.

Residual radioactivity in organs/ tissues/ blood and plasma:

The residual radioactivity was for males and females in all organs and tissues below the limit of quantification. In conclusion, residue data observed were not different from those obtained after single oral administration of U-14C-labelled test substance.

 

METABOLITE PATTERNS

Urine:

There was a presence of up to 8 metabolites (Ul - U7.2) in the 0-48 hour urine samples of rats exposed to a single oral, single intravenous dose of U-14C-labelled test substance, and to a repeated dose (14x) of unlabelled test substance followed by a single dose of U-14C-labelled test substance.

Metabolite U6:

It is by far the most prominent radioactive fraction was U6 identified by co-chromatography on TLC to be test substance itself. Its amount ranged in the various 0-48 hour urine samples from 65.7 to 84.9 % of the radioactivity administered.

Metabolite U1:

Besides the parent molecule, one major, polar metabolite fraction (U1) was found in all urine samples, ranging from 1.3 % (females: single oral high dose) to 9.3 % (males: single intravenous administration). However, when the latter fraction was isolated (males: single oral, high dose) and submitted to TLC in the polar solvent system 79, it was separated into at least 6 metabolites (Ul 1 - Ul.6). Only one minor fraction (U1 2) behaved on TLC identical to reference test substance. All other metabolites were found to be unknown.

Metabolite U2:

Metabolite U2 was an unknown metabolite present in almost all urine samples of the various experiments, ranging from 0.7 to 4.0 %.

Metabolite U3:

Metabolite U3 was a minor metabolite mainly occurring in the 0-48 hour urine samples of rats exposed to a single oral dose of the test substance. Its amount ranged from 0.5 to 1.0 %. It was also detected in urine of males exposed to the repeated oral dose (2.6 %). After isolation by TLC, U3 was identified by co­chromatography was shown.

Metabolite U4:

Metabolite U4 was mainly found in the urine of males exposed to a single low and high dose level, in urine of females treated with the high dose level and in the urine of males intravenously treated with (U-14C)-labelled test substance. It re­presented a minor, unknown metabolite accounting for 0.3 % to 2.9 %.

Metabolite U5:

Metabolite US was a minor, unknown metabolite only found in the urine of males and females exposed to a high, single oral dose of the test substance. The corresponding amounts of U5 were 0.3 % and 0.1 % of the radioactivity administered.

Metabolite U7.1:

Metabolite U7.1 was found in all urine samples of the various groups. It was unknown and its amount ranged from 0.8 to 4.9 %

Metabolite U7.2:

Metabolite U7.2 was only found in the urine of males of the high, single oral dose level study, of females of the intravenous and in the urine of males and females of the repeated oral study. It ranged from 0.6 to 2.8 % of the dose administered. By co-chromatography, the identity of U7.2 was shown. In conclusion, analysis of the various urine samples showed no significant difference of the metabolite patterns in the various groups, i.e. neither the sex and dose level, nor the route of administration influenced the metabolism of the test substance by the rat.

Extractability of radioactivity:

Between 84.2 and 90.5 % of the radioactivity found in faeces was extractable.

Patterns of metabolites in extractables of faeces:

Up to 11 metabolites (F1.1 – F8.2) were found in the extractables of faeces at varying amounts. The most prominent metabolites found in extractables of all treatment groups were F1.1 and F6. Their corresponding amounts ranged from 0.4 to 2.3 % (F1.1) and from 0.7 to 9.6 % (F6 = test substance). All other metabolites ranged, as far as found, from 0.1 to 3.1 % of the dose administered. Based on the TLC­ behaviour in at least two solvent systems, the identity of most faecal metabolites with urinary metabolites U1 - U7.2 could be shown.

Liver and kidney extractability:

The extractability of liver and kidney tissues of rats of groups 1 and 2 (males, females: single oral, high dose level) is discussed. Extractables accounted for 24.0 and 58.9 % of the radioactivity present in the liver of males and females, respectively. These figures corresponded to 0.10 and 0.21 mg parent equivalents per kg of tissue fresh weight.

In kidney of males and females, extractables accounted for 15.4 and 17.7 % of the radioactivity present in the kidney tissues. These figures corresponded to a residual radioactivity level of 0.12 and 0.11 mg/kg.

Non-extractables of males and females were in the liver 0.30 and 0.14 mg/kg and in the kidney 0.68 and 0.50 mg/kg, respectively.

Metabolite pattern:

No analyses could be performed with the extractable radioactivity of kidney of males and females and of the liver of males due to the very low amount of radioactivity present.

Extractables of the liver tissues of female rats were submitted to further analyses. After clean-up, 58.1 % of the radioactivity extracted were recovered corresponding to 34.2 % of the radioactivity present in the liver. This figure corresponded to a residual radioactivity of 0.12 mg/kg.

Analysis by TLC showed the presence of 2 metabolites (L1 and L2) accounting for 0.10 and 0.02 mg/kg. Metabolite fraction L1 proved to be the parent molecule and metabolite L2 was an unknown metabolite.

Conclusions:

The rat metabolism study has shown that the test substance was almost quantitatively absorbed from the gastrointestinal tract by male and female rats. Absorption was rapid after single oral administration, reaching its maximum about 24 hours after the administation. After repeated oral administration, an even more rapid absorption rate was observed, indicating an adaptation of the rats to the test substance as shown by the urinary excretion rates. The major route of excretion was via the urine. Smaller amounts were excreted via the faeces. Elimination of radioactivity was very rapid as shown by the urinary excretion patterns. The half-lives observed were about 24, 8 and less than 8 hours for the single oral, re­peated oral and intravenous route of excretion.
At the low, single oral dose level, residual radioactivity was in almost all organs/tissues, blood and plasma below the limit of quantification. Even when the test substance was repeatedly administered at the low dose level, no radioac­tivity was retained by the animals. At the high target dose level (300 mg/kg), higher levels of residual radioactivity were observed in organs and tissues, indicating that the capacity of the animals to eliminate the test substance de­creased.
Analysis of the metabolite patterns in urine, extractables of faeces and liver showed that mainly the parent molecule was excreted. Besides the parent molecule, a variety of minor metabolites was excreted, i.e. up to at least 12 urinary metabolites and up to at least 8 faecal me­tabolites. No significant difference in the metabolite pat­terns was observed for the various treatment groups, indi­cating that neither the dose level and sex or the routes and rates of administration influenced the metabolism of the test substance by the rat.

Executive summary:

The objectives of the study were to follow the absorption, distribution, excretion and metabolism of the 14C-labelled molecule after single and repeated oral and single intra­venous administration of the test substance at 2 dose levels to male and female rats. For this purpose, the levels of radioactivity appearing in the expired air, urine, faeces, blood and organs/tissues were followed in various separate experiments. Furthermore, the radioactivity appearing in various fractions (liver, kidney, urine and faeces) was characterised by appropriate methods.

The study has shown that the test substance was almost quantitatively absorbed from the gastrointestinal tract of male and female rats. Elimination of radioactivity was rapid and almost quantitative as shown by the urinary/faecal excre­tion rates (the elimination half-life was about 24 hours) and the residual radioactivity levels found in organs/tissues, being after 96 hours below the limit of quantification (LOQ) for the low target dose level (0.5 mg/kg) and in the order of LOQ (spleen, brain, fat, adrenal glands, thyroid gland) up to 0.88 mcg/g (skin, chest) for the high target dose level (300 mg/kg). The latter values indicated a decrease in the elimination capacity of the animals. When the test substance was repeatedly administered at the low target dose level, no radioactivity was retained in organs and tissues of male and female rats. Furthermore, it was shown that repeated oral administration (14x) of the test substance at the low target dose level resulted in a more rapid urinary excretion rate but had no influence on the routes of excretion and on the metabolic capacity of the rat. Analysis of the metabolite patterns in urine, extractables of faeces and extractables of liver showed mainly the presence of the parent molecule, ranging from 65.7 % up to 84.9 % of the dose administered. In faeces, the amount of the test substance ranged from 0.7 % to 9.6 %. Besides the test substance itself, a variety of minor metabolites was ex­creted, i.e. up to at least 12 urinary metabolites totally accounting for 6.6 - 8.1 %, and up to at least 8 faecal meta­bolites totally accounting for 1.2 - 2.9 %. Analysis of ex­tractable radioactivity from liver tissues again showed mainly the presence of the parent molecule besides negligible amounts of one unknown metabolite.

Reference 3

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

Jun 1985 to Apr 1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

metabolism

Qualifier:

no guideline followed

Principles of method if other than guideline:

The objective, of the present study was to isolate the major metabolites excreted by rats after a single oral dose of the test substance, to elucidate their structures by spectroscopic methods and to establish the metabolic pathways of the test substance in rats.

GLP compliance:

not specified

Radiolabelling:

yes

Species:

rat

Strain:

other: Tif: RAI f (SPF)

Sex:

male

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 7 weeks of age
- Weight at study initiation: 200 g
- Housing: Animals were housed individually in metabolism cages.
- Diet: Fasted 18 hr prior to dosing in the morning. Ad libitum. thereafter
- Water: Mains water. Ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 50 ± 10
- Air changes per hour: Not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: Mar 1984 To: Jul 1984

Route of administration:

oral: gavage

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

49 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

10

Control animals:

no

Metabolites identified:

yes

Details on metabolites:

Six urinary metabolites (80 % of the dose) were isolated and identified. The major excretion product identified was the unchanged test substance itself (68 % of the dose). The 5 other urinary metabolites were formed by hydrolysis of the chloroethyl side chain (3,6%), cleavage of the triazine methoxide (2,6 %), cleavage of the sulfonylurea bridge and of the side chain followed by sulfuric acid conjugation (1.7 %), cleavage of the sulfonylurea bridge (0.5 %), hydroxylation of the phenyl ring (0.4 %) and oxidation of the triazine methyl group (3.3 %).

The urinary radioactivity recovered in the 0 - 24, 24 - 48, and 48 - 72 hour periods accounted for 88.6%, 2.6%, and 0.3% of the dose applied. 

 

URINARY METABOLITES
The pooled 0 - 24 h urines (UA), representing 88.6 % of the dose, were used in the following investigations. Two-dimensional TLC in systems SS 1 and SS 2 revealed the presence of 14 metabolite fractions U1 through U14.

The metabolite pattern contained one predominant metabolite (U2, 68 % of the dose) accompanied by 13 minor metabolites (< 3.6 %). High voltage electrophoresis revealed that one major metabolite fraction had a strong acid nature and that most of the others behaved like weak acids. When urine was incubated with S-glucuronidase or with arylsulfatase, no significant change of the metabolite pattern occurred.

Individual metabolites were isolated from the same urine specimen. The urine was chromatographed on LC using LiChroprep RP 18 as stationary phase. The metabolite fractions UA1-1 through UA1-4 were eluted with water and metabolite fraction UA1-5 was eluted with acetonitrile.

Metabolite fraction UA1-2, containing the strong acidic fraction observed by HVE and corresponding to metabolite fraction U14 in the urine pattern, was separated to yield Metabolite 1u. This metabolite was also present in the NMR spectrum probably as a result of hydrolysis during preparation no radioactivity (< 0.1 %) co-chromatographed with the reference material 3 in the urine pattern. Although no cleavage was observed using arylsulfatase in the genuine urine, the purified sulfate metabolite was cleaved by the same enzyme to yield a metabolite fraction co-chromatographing in TLC with reference material 3. Metabolite fraction UA1-5,
containing all the remaining metabolite fractions of the urine pattern, was separated on LiChrosorb RP 18 in two fractions, UA1-52 and UA1-53.

UA1-53 contained exclusively the major metabolite fraction of the urine pattern (U2). This Metabolite U2 was identified by NMR and MS spectroscopy as the test substance itself.

Metabolites 3u, 4u, 5u, and 8u were identified by NMR and MS spectroscopy as reference material 8, 1, 4, 10, respectively; they corresponded to the metabolite frations U11, U8, U1, and U5 of the urine pattern.

Metabolite 7u did not give interpretable NMR and MS spectra, but could be characterised by HPLC and TLC-cochromatography with the reference material 9. It corresponded to metabolite fractions U10.

The structure of metabolite 6u could not be elucidated.

The reference materials 3, 5, 6, 7, 11, and 12 were not found in the urine as checked with TLC (< 0.1 %).

Conclusions:

Most of the administered test substance was absorbed from the intestinal tract into the general circulation and excreted as unchanged test substance. The degradation of the remaining portion proceeded via the following reactions:
(1.) Cleavage of the chloroethyl side chain leading to the phenol derivative reference material 1.
(2.) Oxidation of the triazine methyl group leading to the alcohol derivative reference material 9.
(3.) Cleavage of the triazine methoxy substituent leading to the hydroxy triazine derivative reference material 8. (4.) Hydroxylation of the phenyl ring leading to the phenol derivative reference material 10.
(5.) Cleavage of the sulfonyl urea bridge leading to the sulfonamide derivative reference material 4 and probably reference material 13.
(6.) Further cleavage and conjugation affording the sulfate of reference material 3.
Based on the amount of unchanged test substance excreted, the metabolism of the herbicide in the rat is the same as that found in the goat and in the hen. The cleavage of the sulfonyl urea bridge leading to reference material 4 and its corresponding reference material 13 is a minor pathway in the rat. Hydroxylation of the phenyl ring leading to reference material 10 and 7, the major pathway elucidated in wheat (5.6 % of the applied radioactivity) corresponded to 0.4 % of the dose in the rat.

Executive summary:

The metabolism of the test substance was investigated in rats using U14C-phenyl labelled material.

The urinary radioactivity recovered in the 0 - 24, 24 - 48, and 48 - 72 hour periods accounted for 88.6, 2.6, and 0.3 % of the dose administered, respectively. Six urinary metabolites, representing about 80 % of the dose, were isolated and identified by NMR and MS. The major excretion product isolated and identified was the unchanged test substance itself, which represented 68 % of the dose. The five other urinary metabolites identified, were the phenol 143 (3.6 %, hydrolysis of the chloroethyl side chain), the hydroxy triazine derivative reference material 8 (2,6 %, cleavage of the triazine methoxide), the sulphate of reference material 3 (1.7 %, cleavage of the sulfonylurea bridge and of the side chain followed by sulfuric acid conjugation), the sulfonamide reference substance 4 (0.5 %, cleavage of the sulfonylurea bridge) and reference material 10 (0.4 %, hydroxylation of the phenyl ring). One metabolite was characterised by TLC and HPLC as the al­cohol reference material 9 (3.3 %, oxidation of the triazine methyl group).

Reference 4

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

Mar 1984 to Jul 1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

distribution

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

This study investigated the fate of the (2,6-14 c)-triazine labelled test substance in male and female rats.

GLP compliance:

not specified

Radiolabelling:

yes

Species:

rat

Strain:

other: Tif: RAI f (SPF)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 7 - 8 weeks of age
- Weight at study initiation: 200 g
- Housing: Animals were housed individually in closed all-glass metabolism cages.
- Diet: Fasted 18 hr prior to dosing in the morning. Ad libitum. thereafter
- Water: Mains water. Ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 63 ± 13
- Air changes per hour: Not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: Mar 1984 To: Jul 1984

Route of administration:

oral: gavage

Vehicle:

other:

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- Vehicle: 2.5 % (v/v) Tween 80 (Polysorbate 80), 15.0 % (v/v) Ethanol (absolute), 25.0 % (v/v) Polyethyleneglycol 200 (Carbowax), and 57.5 % (v/v) Phosphate buffer pH 8.0 (0.065 M Na2 HPO4 + 0.004 M KH2 PO4/L)
- 2 mL of the solutions containing 0.051 or 4.87 mg/mL were applied to rats by stomach tube.

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

0.5 mg/kg bw (total dose)

Dose / conc.:

50 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

2

Control animals:

no

Details on absorption:

The test substance is almost completely absorbed from the intestinal tract into the general circulation since 70-97 % of the dose were renally excreted, regardless of the dose applied or of the sex of the animals.

Details on distribution in tissues:

The residual radioactivity, at the 0.5 mg/kg dose level, was under the limit of detection except for 5 values which were smaller or equal to the limit of determination (spleen, kidneys and blood). At the 50 mg/kg dose level, the residues were accordingly higher but still below 0.1 ppm test substance equivalents. Highest values were found in kidneys (0.09 ppm), carcass (0.08 ppm), and blood (0.08 ppm).

Details on excretion:

Within 24 hours 97-100 % of the dose was eliminated, mainly through the kidneys (69-96 %). After 168 hours 70-97 %, 4-31 % and less than 0.3 % of the dose was excreted with urine, faeces and expired air, respectively. Total recoveries of the balance experiment were in the range of 100-102 %. The excretion pattern and the rate of excretion were the same for both sexes and for both dose levels.

Metabolites identified:

no

Details on metabolites:

In the 0-24 h urines, 1 major metabolite fraction (86-95 % of the urine radioactivity) migrating like test substance and 3 minor fractions (up to 6 %) were identified. 

In the pooled 0-24 h faeces, more than 98 % of the faeces radioactivity was extractable. The TLC pattern of the faeces extracts was qualitatively identical to that found with the urine.

DISCUSSION

(2,6-14c-triazine)-test substance, orally applied to male and female rats as single doses of
approximately 0.5 or 50 mg/kg, was almost completely absorbed from the intestinal tract into the general circulation and very rapidly excreted, mainly via the urine. Within 168 hours 70-97 %, 4-31 % and less than 0.3 % of the dose were recovered in urine, faeces and expired air, respectively. Seven days after administration of 0.5 mg/kg, all tissue residues were below or at the limit of determination. The majority of the values were be­low the limit of detection. At the 50 mg/kg dose level the residues in all tissues were accordingly higher, but still below 0.1 ppm test substance equivalents. The TLC pattern of the urine consisted of only 1 major metabolite fraction (86-95 % of the urine radioactivity) migrating like test substance. No significant difference was observed between both sexes and both doses with respect to rate and pattern of excretion. The metabolite patterns in faeces and urine were qualitatively the same. The data obtained (2,6-14c-triazine)-test substance in the rat are essentially the same as those reported earlier with (u-14c-phenyl)-substance. Therefore, the bridge between the s-triazine and the phenyl ring was not cleaved to a significant extent. There is strong evidence that the major portion of the test substance applied is excreted unchanged.

 

Table 1. Excretion of radioactivity by rats after a single oral dose of (2,6-14c-triazine)-test substance (in % of dose).

	Low Dose				High Dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2
Dose (mg/kg)	0.65	0.54	0.52	0.47	59.1	51.3	49.2	58.0
Faeces
0-24 hr	11.58	2.95	4.21	3.42	7.98	29.24	5.80	4.78
24-48 hr	0.48	0.78	0.62	0.91	0.83	1.00	1.85	1.05
48-72 hr	0.11	0.08	0.10	0.18	0.12	0.11	0.12	0.28
72-168 hr	0.17	0.06	0.04	0.44	0.18	0.14	0.07	0.24
Sub-total	12.35	3.88	4.99	4.95	9.11	30.50	7.84	6.35
Urine
0-24 hr	88.16	95.91	95.41	93.02	90.74	69.04	91.22	93.83
24-48 hr	0.76	0.74	0.66	1.30	1.37	0.95	2.41	1.28
48-72 hr	0.26	0.18	0.15	0.24	0.17	0.16	0.07	0.29
72-168 hr	0.29	0.11	0.13	0.20	0.13	0.19	0.07	0.26
Sub-total	89.48	96.93	96.35	94.76	92.40	70.34	93.78	95.67
Expired air
0-24 hr	0.04	0.06	0.15	0.11	0.04	0.03	0.02	0.04
24-48 hr	0.01	0.01	0.02	0.02	0.01	0.01	0.01	0.01
48-168 hr	0.00	0.00	0.03	0.01	0.00	0.00	0.00	0.00
Sub-total	0.05	0.07	0.20	0.14	0.05	0.04	0.03	0.05
Total excretion	101.87	100.88	101.53	99.86	101.56	100.87	101.65	102.07
Tissue residues1)	0.02	0.02	0.02	0.02	0.03	0.02	0.02	0.02
Cage wash	0.07	0.03	0.12	0.52	0.22	0.06	0.19	0.13
Total recovery	101.96	100.93	101.67	100.40	101.81	100.95	101.86	102.23

1) Calculated from Table III assuming that fat, blood and muscle
represent 11 %, 5.9 % and 45 % of the body weight, respectively (5,6,7) but without carcass values.

 

Table 2. Residual radioactivity in selected tissues of rats seven days after a single oral dose of (2,6-14c-triazine)-test substance (in ppm test substance equivalents)

	Low Dose				High Dose				Low dose	High dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2	LQ1 (ppm)
Dose (mg/kg)	0.65	0.54	0.52	0.47	59.1	51.3	49.2	58.0
Tissue
Spleen	<LD	<LQ	<LQ	<LQ	~LQ	<LQ	~LQ	<LQ	0.002	0.020
Liver	<LD	<LD	<LD	<LD	0.031	0.031	0.013	0.021	0.001	0.006
Fat	<LD	<LD	<LD	<LD	~LD	<LD	<LD	<LD	0.003	0.033
Kidneys	<LD	<LD	LQ	<LD	0.071	0.058	0.054	0.091	0.001	0.009
Muscles	<LD	<LD	<LD	<LD	0.011	0.011	~LQ	0.01	0.001	0.005
Blood	<LD	<LD	<LD	LQ	0.072	0.040	0.051	0.076	0.001	0.007
Brain	<LD	<LD	<LD	<LD	<LD	<LD	<LD	LD	0.001	0.012
Heart	<LD	<LD	<LD	<LD	<LQ	<LQ	~LD	<LQ	0.002	0.025
Lungs	<LD	<LD	<LD	<LD	0.022	~LQ	0.021	0.024	0.001	0.012
Testes	<LD	<LD	-	-	<LQ	<LQ	-	-	0.001	0.008
Ovaries	-	-	<LD	<LD	-	-	<LD	<LD	0.005	0.051
Carcass	<LD	<LD	<LD	<LD	0.082	0.052	0.019	0.035	0.001	0.005

1 LQ= Limit of determination, LD= Limit of detection, LD= 0.33 LQ. Values were calculated according to CURRIE.

 

Table 3. Distribution of urinary metabolites (% of the urine radioactivity) of (2,6-14c-triazine)-test substance.

	Low Dose				High Dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2
Dose (mg/kg)	0.65	0.54	0.52	0.47	59.1	51.3	49.2	58.0
Urine 0-24 hrs
Met. Fr.U1	93	93	94	95	86	87	92	88
Met. Fr.U2	4	3	3	2	6	5	3	5
Met. Fr.U3	2	2	1	1	4	4	3	4
Met. Fr.U4	2	2	3	2	4	4	2	3

Conclusions:

The test substance, orally applied to male and female rats at single doses of approximately 0.5 or 50 mg/kg, was almost completely absorbed and mainly excreted via the urine. The TLC pattern of the urine consisted of only one major metabolite fraction (86-95 % of the urine radio­activity) migrating like the test substance. There is strong evidence that the major portion of the test substance applied is excreted unchanged.

Executive summary:

The fate of the test substance was investi­gated in male and female rats using (2,6-14c)-triazine labelled material.

Single oral doses of approximately 0.5 or 50 mg/kg were readily absorbed from the intestinal tract into the general circulation and very rapidly excreted. From 97-100 % of the doses ad­ministered were eliminated within 24 hours, mainly through the kidneys. After 168 hours 70-97 %, 4-31 % and less than 0.3 % were ex­creted with urine, faeces and expired air, respectively. Total recoveries of the balance experiment were in the range of 100-102 %. The excretion pattern and the rate of excretion were similar for both sexes and both dose levels. The residual radioactivity was determined 168 hours after dosing in spleen, liver, fat, kidneys, muscles, brain, blood, lungs, heart, testes, ovaries and in the remaining carcass. At the 0.5 mg/kg dose level (corresponding to 5 ppm test substance in the 0-24 hours food) the residual radioactivity was below the limit of detection (LD) except for five values which were still below or equal to the limit of determination (LQ). At the 50 mg/kg dose level residues were accordingly higher but still below 0.1 ppm. The urine was analysed by TLC showing only one major meta­bolite fraction (86-95 % of the urine radioactivity) migrating like the test substance. More than 98 % of the faeces radioactivity was extractable. The major metabolite fraction of the faeces extracts (about 73 % of the faeces radioactivity) behaved like that present in the urine. The data reported here are essentially the same as those ob­tained in the identically designed experiment using (u-14c-phenyl)-test substance. Therefore, the cleavage of the bridge between the s-triazine and the phenyl ring during the passage of the test substance through the animal did not occur to a significant extent resulting in the major portion of the test substance applied to be excreted unchanged.

Reference 5

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

Jan 1984 to Mar 1984

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Objective of study:

distribution

excretion

Qualifier:

no guideline followed

Principles of method if other than guideline:

The fate of the test substance was investigated in male and female rats using (U-14C)-phenyl labelled material in male and female rats.

GLP compliance:

not specified

Radiolabelling:

yes

Species:

rat

Strain:

other: Tif: RAI f (SPF)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 7 - 8 weeks of age
- Weight at study initiation: 200 g
- Housing: Animals were housed individually in closed all-glass metabolism cages.
- Diet: Fasted 18 hr prior to dosing in the morning. Ad libitum
- Water: Mains water. Ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2
- Humidity (%): 65 ± 10
- Air changes per hour: Not specified
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: Jan 1984 To: Mar 1984

Route of administration:

oral: gavage

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- Vehicle: 2.5 % (v/v) Tween 80 (Polysorbate 80), 15.0 % (v/v) Ethanol (absolute), 25.0 % (v/v) Polyethyleneglycol 200 (Carbowax), and 57.5 % (v/v) Phosphate buffer pH 8.0 (0.065 M Na2 HPO4 + 0.004 M KH2 PO4/L)
- Dosing volume: 2 mL of the solutions containing 0.052 mg/mL or 5.02 mg/mL were applied to rats by a stomach tube.

Duration and frequency of treatment / exposure:

Single dose

Dose / conc.:

0.1 mg/kg bw (total dose)

Dose / conc.:

50 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

2

Control animals:

no

Details on absorption:

The test substance was almost completely ab­sorbed from the intestinal tract into the general circulation since 83-99 % of the dose was renally excreted.

Details on distribution in tissues:

The residual radioactivity, at the 0.5 mg/kg dose level, was under the limit of detection except for two values which were below or equal to the limit of determination (ovary and blood). At the 50 mg/kg dose level, the residues were accordingly higher but most of the values were below 0.1 ppm test substance equivalents, except values found in the ovaries (0.1-0.3 ppm) and in 1 sample of blood (0.2 ppm).

Details on excretion:

Within 24 hours 92-100 % of the dose was eliminated, mainly through the kidneys (80-99 %). After 168 hours 83-99 %, 1-16 % and less than 0.1 % of the dose was excreted with urine, faeces and expired air, respectively. The excretion pattern and the rate of excretion were the same for both sexes and for both dose levels. Total recoveries of the balance experiment were in the range of 98-102 %.

Metabolites identified:

yes

Details on metabolites:

In the 0-24 hour urines, one major metabolite fraction was identified (70-95 % of the urine radioactivity) migrating like the test substance and 3 minor fractions (up to 13 %). 

In the pooled 0-24 h faeces, more than 93 % of the faeces radioactivity was extractable. The TLC pattern of the faeces extracts was qualitatively identical to that found with the urine.

DISCUSSION

Radiolabelled test material orally applied to male and female rats at single doses of approximately 0.5 or 50 mg/kg, was almost completely absorbed from the intestinal tract into the general circulation and very rapidly excreted, mainly via the urine. Within 168 h 83-99 %, 1-16 % and less than 0.1 % of the dose were recovered in urine, faeces and expired air, respectively. Seven days after administration of 0.5 mg/kg, all tissue residues were below or at the limit of determination. The majority of the values were even below the limit of detection. At the 50 mg/kg dose level the residues in all tissues were accordingly higher but residues above 0.1 ppm equivalents were found only in the ovaries (0.1 and 0.3 ppm) and in one sample of blood (0.2 ppm). The TLC pattern of the urine consisted of only one major metabolite fraction (70-95 % of the urine radioactivity) migrating like. The major metabolite excreted with the faeces behaved like that present in the urine, i.e. cochromatographing with the test substance. No significant difference was observed between both sexes and both doses with respect to rate and pattern of excretion. The metabolite pattern in faeces and urine were qualitatively the same. There is strong evidence that the major portion of the test substance applied is excreted unchanged.

 

Table 1. Excretion of radioactivity by rats after a single oral dose of(u-14c-phenyl)-test substance (in % of dose).

	Low Dose				High Dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2
Dose (mg/kg)	0.54	0.56	0.54	0.57	52.9	51.3	54.64	57.1
Faeces
0-24 hr	2.76	4.72	1.17	9.46	5.98	12.48	6.65	3.82
24-48 hr	0.28	0.67	0.15	1.58	0.56	2.68	1.01	0.80
48-72 hr	0.07	0.06	0.02	0.36	0.08	0.29	0.23	0.12
72-168 hr	0.11	0.08	0.02	0.02	0.17	0.23	0.30	0.12
Sub-total	3.23	5.53	1.36	11.42	6.79	15.69	8.20	4.86
Urine
0-24 hr	94.57	94.23	98.60	86.78	91.80	79.62	92.01	95.43
24-48 hr	0.30	0.55	0.14	0.24	0.82	2.59	0.75	1.66
48-72 hr	0.08	0.33	0.02	0.04	0.17	0.20	0.08	0.11
72-168 hr	0.10	0.13	0.01	0.04	0.11	0.14	0.10	0.11
Sub-total	95.06	95.14	98.79	87.10	92.89	82.55	92.94	97.31
Expired air
0-24 hr	0.01	0.01	0.01	0.01	0.01	0.01	0.01	0.1
24-48 hr	0.01	0.00	0.00	0.01	0.00	0.00	0.00	0.00
48-168 hr	0.00	0.00	0.00	0.00	0.00	0.00	0.00	0.00
Sub-total	0.02	0.01	0.01	0.02	0.01	0.01	0.01	0.01
Total excretion	98.31	100.68	100.16	98.54	99.69	98.25	101.15	102.18
Tissue residues1)	0.03	0.01	0.01	0.01	0.02	0.08	0.01	0.03
Cage wash	0.04	0.05	0.01	0.27	0.02	0.02	0.12	0.08
Total recovery	98.38	100.74	100.18	98.82	99.73	98.35	101.28	102.29

1) Calculated from Table III assuming that fat, blood and muscle represent 11%, 5.9% and 45% of the body weight, respectively (4,5,6) but without carcass values.

 

Table 2. Residual radioactivity in selected tissues of rats seven days after a single oral dose of (u-14c-phenyl)-test substance (in ppm test substance equivalents)

	Low Dose				High Dose				Low dose	High dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2	LQ1 (ppm)
Dose (mg/kg)	0.54	0.56	0.54	0.57	52.9	51.3	54.64	57.1
Tissue
Spleen	<LD	<LD	<LD	<LD	<LD	0.030	~LD	<LD	0.002	0.021
Liver	<LD	<LD	<LD	<LD	0.035	0.135	0.016	0.025	0.001	0.008
Fat	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	0.006	0.066
Kidneys	<LD	<LD	<LD	<LD	0.048	0.125	0.042	0.075	0.001	0.010
Muscles	<LD	<LD	<LD	<LD	~LD	0.022	~LD	~LD	0.001	0.011
Blood	<LD	<LD	<LD	<LD	0.034	0.160	0.026	0.068	0.001	0.008
Brain	<LD	<LD	<LD	<LD	<LD	<LD	<LD	<LD	0.002	0.016
Heart	<LD	<LD	<LD	<LD	~LQ	0.056	<LQ	0.022	0.002	0.015
Lungs	<LD	<LD	<LD	<LD	~LQ	0.073	<LQ	0.029	0.002	0.017
Testes	<LD	<LD	-	-	0.011	0.037	-	-	0.001	0.008
Ovaries	-	-	<LD	<LD	-	-	0.131	0.273	0.007	0.101
Carcass	<LD	<LD	<LD	<LD	0.051	0.073	0.040	0.023	0.001	0.009

1 LQ= Limit of determination, LD= Limit of detection, LD= 0.33 LQ. Values were calculated according to CURRIE.

 

Table 3. Distribution of urinary metabolites (% of the urine radioactivity) of (u-14c -phenyl)-test substance.

	Low Dose				High Dose
	Male 1	Male 2	Female 1	Female 2	Male 1	Male 2	Female 1	Female 2
Dose (mg/kg)	0.54	0.56	0.54	0.57	52.9	51.3	54.64	57.1
Urine 0-24 hrs
Met. Fr.U1	92	93	95	95	92	70	93	90
Met. Fr.U2	~4	~3	~2	~2	4	13	4	~6
Met. Fr.U3	< 2	< 2	<2	<2	2	~7	<2	<2
Met. Fr.U4	<2	<2	<2	<2	~2	~9	~2	~2

 

Conclusions:

The test substance, orally applied to male and female rats at single doses of approximately 0.5 or 50 mg/kg, was almost completely absorbed and mainly excreted via the urine. The TLC pattern of the urine consisted of only one major metabolite fraction (70-95 % of the urine radio­activity) migrating like the test substance. There is strong evidence that the major portion of the test substance applied is excreted unchanged.

Executive summary:

The fate of the test substance was investi­gated in male and female rats using (u-14c)-phenyl labelled material.

Single oral doses of approximately 0.5 or 50 mg/kg bw were readily absorbed from the intestinal tract into the general circulation and very rapidly excreted. More than 92 % of the doses were eliminated within 24 hours, mainly through the kidney. After 168 hours 83-99 %, 1-16 % and less than 0.1 % were excrete with urine, faeces and expired air, respectively. Total recovery of the balance experiment were in the range of 98-102 %. The excretion pattern and the rate of excretion were similar for both sexes and both dose levels. The residual radioactivity was determined 168 hours after dosing in spleen, liver, fat, kidneys, muscles, brain, blood, lungs, heart, testes, ovaries and in the remaining carcass. At the 0.5 mg/kg bw dose level (corresponding to 5 ppm in the 0 - 24 hours food) the residual radioactivity was below the limit of detection (L0) except for two values which were still below or equal to the limit of determination (LQ). At the 50 mg/kg bw dose level residues were accordingly higher but most of the values were still below 0.1 ppm equivalents. The urine was analysed, showing only one major metabolite fraction (70-95 % of the urine radioactivity) migrating like test substance. More than 93 % of the faeces radioactivity was extractable. The major metabolite fraction of the faeces extracts (about 72 % of the faeces radioactivity) behaved like that present in the urine. No significant difference was observed between both sexes and both doses with respect to rate and pattern of excretion. The metabolite pattern in faeces and urine were qualitatively the same. There is strong evidence that the major portion of the test substance applied is excreted unchanged.

Description of key information

- Oral absorption: the test substance was rapidly and almost completely absorbed within 24 hours (Hagenbuch 1985a, 1985b, and 1986, Capps 1987)

- Metabolism: Only a small portion of the parent compound was metabolised. The major pathways steps were demethylation of the methoxy group, phenol ring hydroxylation, methyl hydroxylation, whereas the sulfonylurea bridge was not cleaved to a significant extent. (Van Dijk 1987, Hagenbuch 1986)

- Excretion: The test substance is rapidly excreted mainly via the urine and to a lesser extend the faeces and expired air (Hagenbuch 1985a, 1985b, and 1986, Capps 1987)

- Dermal absorption: 19 % and 20% (1 g/L and 0.1 g/L spray concentration for 24 h exposure), in vivo absorption study in rats (Murphy 1990).

- Inhalation absorption: Based on the lack of information on inhalation absorption, the default absorption value of 100% from the REACH guidance (Chapter 8, R.8.4.2) is used for DNEL derivation.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Absorption rate - oral (%):

100

Absorption rate - inhalation (%):

100

Additional information

Oral absorption: In rats, a single dose of the test substance was rapidly absorbed in the intestinal tract regardless of the label (U14-C-phenyl or 14C-triazine label) and was very rapidly excreted, mainly via the urine (> 70 % within 168 hours) (Hagenbuch 1985a, 1985b, 1986, Van Dijk 1987). A small percentage of the dose was recovered in the faeces and expired air (> 1 and > 0.1 %, respectively). Additionally, the test substance was readily absorbed following repeated dose.

Dermal absorption: The test substance was partially dermally absorbed in rats at doses of 0.02 or 0.2 mg/rat, 20% or 19 % of diluted product, respectively (Murphy 1990). In vitro, the test substance was dermally absorbed to a lesser extent of 0.04, 1.8, and 4.4 %, for the 33.3 % w/w slurry, 1/444 w/w dilution, and 1/888 w/w dilution, respectively (Johnson 2012).

Distribution: In rats, following a single oral dose of 0.5 or 300 mg/kg, tissue residues were <0.1 ppm at the top dose and below the level of determination at the low dose after 168 hours (Hagenbuch 1985a, Kl2). In rats, the residual radioactivity levels found in organs/tissues after 96 hours were below the LOQ for the low dose level (0.5 mg/kg) and up to 0.88 mcg/g for the high target dose level (300 mg/kg) (Van Dijk 1987). In another study, the total radioactive residues in all tissues were below 0.1 ppm (0.5 and 50 mg/kg dose levels) (Capps 1987).

Metabolism: The major compound in the urine was the parent compound amounting to 68 - 85 % of the applied dose. The major pathways steps were demethylation of the methoxy group, phenol ring hydroxylation, methyl hydroxylation, whereas the sulfonylurea bridge was not cleaved to a significant extent. No difference in the metabolite pattern was noted between dose levels and sexes (Hagenbuch, 1986).

Excretion: Within 24 h, 74-98% of the dose applied was excreted in urine and 2-14% in faeces. The excretion pattern and the rate of excretion were similar for both sexes and for high and low dose levels. Excretion was faster after repeated administration compared to single administration. The half-lives were approximately 24, 8 and less than 8 hours following single oral, repeated oral and intravenous administration of the compound, respectively.