N-ethyl-o(or p)-toluenesulphonamide

Endpoint summary

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

July 2006- July 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed according to guideline and under GLP.

Objective of study:

toxicokinetics

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

GLP compliance:

yes

Radiolabelling:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
ADME and TK groups (1-6) : Rat: Wistar, Crl:WI (outbred, SPF quality).Bile-cannulation group (7) : Rat: Wistar HsdCpb : WU.
- Source: ADME and TK groups (1-6) : Charles River Deutschland, Sulzfeld, Germany. Bile-cannulation group (7) : Harlan, Horst, The Netherlands
- Age at study initiation: Young adult animals of 10-12 weeks old were used.
- Weight at study initiation: body weights ≥ 179 gram
- Fasting period before study: No
- Housing:
ADME and TK groups (1-6):
Upon receipt from the supplier animals were group-housed in Macrolon cages (type MIV, height 18 cm) containing sterilized sawdust bedding (Woody-Clean type 314; Tecnilab-BMI BV, Someren, The Netherlands). Certificates of analysis were examined and then retained in the NOTOX archives. In
addition, paper was provided as nest material (Enviro-dri, Tecnilab-BMI BV, Someren, The Netherlands). Animals of groups 3 and 6 remained housed in these cages during the pre-treatment. Following administration of the radioactive dose and 24 hours prior to dosing, rats from ADME groups (1-3) were individually housed in stainless steel metabolism cages (LxWxH = 18.5x19x20 cm). Animals of TK groups (4-6) were housed individually in
Macrolon cages (LxWxH: 30x1 9x1 9 cm), equipped with a bottom grid and paper bedding. Surplus animals were kept in the Macrolon cages.
Bile-cannulation group (7):
Upon receipt from the IMTC animals were individually housed in Macrolon plastic cages (type Mlll height 15 cm) containing sterilized sawdust bedding (Woody-Clean type 314; Tecnilab-BMI BV, Someren, The Netherlands) and tissues. Certificates of analysis of the bedding were examined for contaminants and archived. After the acclimatization period, animals were individually housed in stainless steel metabolism cages (LxWxH = 18.5x19x20 cm). Two surplus animals were kept in the Macrolon cages.
- Individual metabolism cages: yes
- Diet (e.g. ad libitum): ad libitum access to pelleted rodent diet (SM RIM-Z from SSNIFFB Spezialdiaten GmbH, Soest, Germany)
- Water (e.g. ad libitum): ad libitum
- Acclimation period: At least 5 days under laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.0-22.0
- Humidity (%): 45-90
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES:
Grow 1
Start of administration: 18 July 2006
Termination: 21 July 2006

Group 2
Start of administration: 24 July 2006
Termination: 27 July 2006

Group 3
Start of administration "cold": 21 July 2006
Start of administration C14: 31 July 2006
Termination: 03 August 2006

Group 4
Start of administration: 18 July 2006
Blood sampling for toxicokinetics: 18, 19, 20 and 21 July 2006
Termination: 21 July 2006

Group 5
Start of administration: 24 July 2006
Blood sampling for toxicokinetics: 24, 25, 26 and 27 July
Termination: 27 July 2006

Group 6
Start of administration "cold": 21 July 2006
Start of administration C14: 31 July 2006
Blood sampling for toxicokinetics: 31 July, 01, 02 and 03 August 2006
Termination: 03 August 2006

Group 7
Administration: 28 August 2006
Termination: 30 and 31 August 2006

Experimental completion date: 30 and 31 August 2006

Route of administration:

oral: gavage

Vehicle:

water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
A weighed amount of unlabeled Chloramine T was placed into an empty glass container. A measured amount of a stock solution of labelled Chloramine T in Milli-Q water and additional Milli-Q water was added to obtain the desired concentration and specific activity. All radiolabelled solutions were prepared immediately prior to dosing and stored at ambient temperature, for a maximum of 4 hours. Before and during the treatment procedure the
treatment solutions were kept on a magnetic stirring device.

The unlabelled Chloramine T formulation used for pre-treatment was prepared immediately prior to dosing and stored at ambient temperature for a maximum of 4 hours.


VEHICLE
Milli-Q water for the labelled formulations.

Milli-U water for the non-labelled Chloramine T formulation (used for the 10 days pre-dosing of group 3 and 6 animals).
- Justification for use and choice of vehicle (if other than water): not applicable
- Concentration in vehicle: Stock solutions of [ring-U-14C]Chloramine T were prepared in Milli-Q water. Four dosing formulations were prepared with the following nominal concentration and specific activity:
Group 1 and 4: 4 mg/ml with a specific activity of 0.5 MBq/mg
Group 2 and 5: 40 mg/ml with a specific activity of 0.05 MBq/mg
Group 3 and 6: 4 mg/ml with a specific activity of 0.5 MBq/mg
Group 7: 4 mg/ml with a specific activity of 0.3 MBq/mg
- Amount of vehicle (if gavage): 5ml/Kg
- Lot/batch no. (if required): not applicable
- Purity: not applicable


HOMOGENEITY AND STABILITY OF TEST MATERIAL:
Before and after dose administration, the homogeneity and radioactivity concentration of each formulation was verified by radioanalysis. For this purpose, weighed aliquots of 25 µl were taken from the top, middle and bottom of the mixture and analyzed by liquid scintillation counting (LSC) using Ultima Gold (Packard) scintillation cocktail.

Duration and frequency of treatment / exposure:

Single dose for groups 1, 2, 4, 5 and 7. In the case of groups 3 and 6, a single oral dose of labelled Chloramine T was administered following pre-treatment with non-labelled Chloramine T for 10 days.

Remarks:

Doses / Concentrations:
ADME and TK groups:
Groups 1 and 4: low-dose of 20 mg/kg bw.
Groups 2 and 5: high-dose of 200 mg/kg bw.
Groups 3 and 6: low-dose of 20 mg/kg bw.
Each oral dose of [ring-U-14C]Chloramine T contained approximately 10 MBq/kg bw of radioactivity.
Bile-cannulation group 7: a single low-dose of 20 mg/kg bw, containing approximately 6 MBq/kg bw of radioactivity.

No. of animals per sex per dose / concentration:

4/sex/dose level (ADME groups: 1-3)
3/sex/dose level (TK groups: 4-6)
4/sex/dose level (bile-cannulation group: 7)

Control animals:

no

Details on study design:

- Dose selection rationale:
Dose levels were chosen based on available acute toxicity data and a 90-d dietary study in rats. The LD50 of a 10% solution of Chloramine T was 935 mg/kg bw. The effect level in the 90-d study was > 150 mg/kg bw/day and the NOAEL was 15 mg/kg bw/day (studies avaialble at that time from 1971). Based on this, the low-dose level in the ADME study was chosen as 20 mg/kg bw and the high-dose level as 200 mg/kg bw. Furthermore, the dose levels were a factor 10 apart, which allowed the investigation of differences in the ADME patterns and bioavailability as a function of dose level.
- Rationale for animal assignment (if not random): random

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled : urine, faeces, blood, plasma, serum, cage washes, bile, bone, Heart, Lung, Spleen, Gonads (testes, seminal vesicles, prostate, uterus, ovaries: all collected separately), Abdominal fat, Muscle, Adrenals, Thymus, Thyroid, Liver, Kidney, Brain, Bone
GI tract (including contents), Skin (shaved), Residual carcass
- Time and frequency of sampling:
For the animals in ADME and bile-cannulation groups (1-3 and 7) bile (group 7 only), urine and faeces were collected over the following time intervals: 0-8, 8-24, 24-48 and 48-72 hours after the dosing with [ring-U-14C]Chloramine T. Urine, bile and faeces were freeze-trapped to avoid atmospheric oxidation, evaporation and bacterial degradation. Urine, bile and faeces samples were stored at ≤ 75°C prior to analysis. At termination, the interior of the metabolism cages was rinsed with methanol/water (50/50). The cage rinse was weighed and stored at 5 -10°C until analyzed. Based on the results of pilot study 462105 (non-GLP), volatiles were not sampled.
For the animals in TK groups (4-6), blood samples from the tail vein of each rat were sampled at the following time-points after dose administration:
15, 30 min, 1 , 2, 4, 8, 24, 48 and 72 hours (max. deviation 1 minute). The amount of sample collected was approximately 300 µl per sampling event. At approximately half an hour prior to blood sampling the animals of groups 4, 5 and 6 were transferred to an incubator set at 40°C where they remained until the time of blood sampling. At the times of blood sampling, the rats were maintained in restrainers. At other times they were housed individually in Macrolon cages and allowed food and water ad libitum. After the final blood sample was obtained, the animals were euthanized by a C02/02 procedure and the carcasses disposed of as radioactive waste. Sampled blood was transferred into tubes containing Liheparin and centrifuged to obtain the plasma which was stored at ≤ -75°C prior to analysis.
At the designated time of euthanasia each animal in ADME groups (1-3) was deeply anaesthetised using isoflurane vapour. The thorax and abdomen were opened with a midline incision. By means of an aorta punction the maximum possible amount of blood was withdrawn, causing the animals to be exsanguinated. Care was taken to avoid contamination of other organs with blood. Sampled blood was transferred into tared tubes containing Li-heparin and weighed. A weighed subsample of approximately 1 ml of the heparinised blood was removed for total 14C analysis. The remaining blood was centrifuged to obtain the plasma which was weighed and stored at ≤ -75°C. The organs listed above were harvested.
Samples from muscle and bone were collected from the left hind leg. The weight of each tissue and blood sample were recorded at the time it was harvested. Tissues and carcass were stored at ≤ -10°C prior to analysis.
At the designated time of euthanasia all animals of bile-cannulation group 7 were euthanized using an oxygen/carbon dioxide procedure. The carcass and GI tract (including contents) were stored at ≤ -10°C prior to analysis.
- Other: None


METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine, faeces
- Time and frequency of sampling:
Urine:
ADME Groups (1 -3): 0-8 and 8-24 h
Faeces:
ADME Groups (1-3): 8-24 h
- From how many animals: the samples were pooled in order to obtain one urine and faeces sample per group per sex. That is 4 animals per group per sex
- Method type(s) for identification: LC-RAD-PDA-MS and NMR
- Limits of detection and quantification: see: additional information on materials and methods
- Other: None

Statistics:

Mass balance:
The mean and standard deviation was used to characterize the data, where appropriate (i.e. radioactivity measurement, concentration, etc). Concentrations of radioactivity in blood, plasma and tissues were calculated as mg equivalents of Chloramine T /kg of sample, based on the specific activity of the [ring-U-14C]Chloramine T formulation. The total amounts of radioactivity in tissue and excreta samples were calculated as a percentage of the administered radioactivity by the following formula:
Amount excreted (% of administered dose) = (radioactivity matrix (Bq/total matrix)/dose (Bq/animal))* 100%
and a 14C mass balance were presented. A mean recovery of I00 ± 10% of the administered radioactive dose was achieved. For ADME groups (1-3) and the bile-cannulation group (7): Cumulative amounts (expressed as percentage of the dose) excreted in urine, bile and faeces were calculated by summation of the amounts excreted in the individual urine, bile and faeces samples per collection period, respectively.
The oral absorption was calculated by summing the amounts of radioactivity in urine, bile (group 7), cage-wash and tissues and organs.

Kinetic calculations:
Concentrations of radioactivity in plasma were calculated as mg equivalents of Chloramine T /kg of sample, based on the specific activity of the [ring-U-14C]Chloramine T formulation. For each group, all toxicokinetic parameters were calculated from the curves constructed from the mean values at each time point using the WinNonlin 4.0.1 program. Non-compartmental analysis was applied.

Details on absorption:

The absorption of Chloramine T equivalents was high, 92% and 91% for the males and females in group 1 respectively, 83% in the males and females of group 2, and 89% and 85% of the males and females in group 3 respectively. No significant differences were noted in absorption between low and high dosing and single and repeated dosing. The absorption in the bile-cannulation group was lower than in the ADME groups, 75% in both sexes. Urinary excretion of radioactivity in the bile-cannulated animals was about 11 to 23% lower than in the ADME groups and excretion in the faeces was about 7 to 15% higher. The reason for this is unclear: animals in the bile-cannulation group showed more clinical signs and can perhaps be viewed as in worse condition than the other animals, possible influencing absorption. Another explanation can be that the absence of bile influences the absorption of the compound. No sex-differences were noted in oral absorption in any of the groups.

Details on distribution in tissues:

The average total remaining radioactivity in blood, carcass plus tissues in the ADME groups was between 0.6 and 1.4% of the administered dose in all groups. The major part of it was present in carcass, i.e. between 0.4 and 1.1 % of the administered dose in all groups. In the bile cannulation group, the average total remaining radioactivity in carcass and GI tract was less than 1 % of the administered dose.
The residual concentrations of Chloramine T equivalents in group 1 and 3 were in the same
range; concentrations in group 2 (high dose group) were approximately one order of magnitude
higher than in the low dose groups.
The tissue concentration equivalents of Chloramine T were below the concentration observed in blood in all groups. Highest concentrations in tissueslorgans were observed in liver and kidney, which is indicative of the extensive metabolism and excretion of the substance. This indicates that Chloramine T shows no potential for accumulation.

Details on excretion:

Excretion via urine is an important route of elimination for Chloramine T, after oral absorption. Urinary excretion accounted for 90% for males and 88% for females after low dosing, 81% for males and 79% for females after high dosing and 87% for males and 78% for females after repeated dosing. In the bile-cannulation group urinary excretion accounted for 67% for the males and 66% for the females after oral dosing. The rate of excretion was similar for all ADME groups with the bulk of radioactivity excreted during the first 8h. The excretion rate of radioactivity in the urine in the bile-cannulation group was slightly lower than in the ADME groups in the first time intervals.

Excretion via faeces was a minor route of excretion for Chloramine T equivalents. Faecal excretion accounted for 10% for the males and females after
low dosing, 13% for the males and females after high dosing and 13% for the males and 16% for the females after repeated dosing, in the ADME groups. In the bile-cannulation group faecal excretion accounted for 25% for the males and 23% for the females. The excretion of radioactivity in faeces was delayed compared to urine, with significant excretion not until after 8 h post-dose. The highest amount of radioactivity was excreted in the
8-24 h interval.

Excretion via the bile is only a minor route of excretion for for Chloramine T equivalents, after oral absorption. Biliary excretion accounted for 6% of the administered dose in males and 4% in females. Excretion took place mainly in the first interval, from 0-8 hours after dosing.

The results show that 95-101% of the administered dose was excreted during the study period (mean values). Excretion was quick and almost complete within 24h post-dose.

Toxicokinetic parameters:

other: see table below: remarks on results including tables and figures

Metabolites identified:

yes

Details on metabolites:

LC-RAD-PDA-MS metabolite analysis
An overview of the data obtained in the metabolite identification part of this study indicated that a wide variety of compounds have been found that may be metabolites of p-TSA, which is formed from the chemically instable Chloramine T. Most of these compounds could not be identified.
In the urine samples, one major radioactivity peak was observed. Despite analysis in negative and positive modes no identification of potential metabolites was possible. Despite the large numbers of potential metabolites that were found with LC-MS analysis in the faeces extracts of the Wistar rats, only two potential metabolites were identified, i.e. p-sulfonicbenzoic acid and an aromatic methoxy compound. In the radioactivity chromatograms 3 major peaks were observed. None of the identified metabolites did correspond to these peaks.

NMR metabolite analysis
The urine of group 2 males was used for analysis. The unknown metabolite, representing approx. 80% of the radioactivity in the chromatogram, was isolated by prep HPLC. Analysis of 1H and 13C NMR indicated that the isolated metabolite was a para-substituted benzene with COOH and SO2NH2 substituents, and identified as 4-Sulfamoyl benzoic acid. In addition, mass spectrometric analysis was carried out which was consistent with the molecule being 4-Sulfamoyl benzoic acid.

Dose verification: The radioactive dose administered to each animal was calculated from the weight of the dose administered and the mean radioactive concentration of the formulations. Animals in ADME and TK groups (1,-6) received average effective radioactive doses ranging between 10.2 and 11.1 MBq/kg bw. Animals in the bile-cannulation group (7) received an average effective radioactive dose of 6.3 MBq/kg bw. Animals in the low-dose groups received an average dose of Chloramine T between 20.1 and 20.6 mg/kg bw. Animals in the high-dose groups received an average dose of Chloramine T between 190.8 and 205.4 mg/kg bw. All the doses were sufficiently close to the target doses in order to fulfill the study's objectives. Toxicokinetic parameters

		group 4 20 mg/kg bw single oral dose		group 5200 mg/kg bw single oral dose		group 620 mg/kg bw repeated oral dose
parameters		m	f	m	f	m	f
Tmax	hr	0.5	0.5	1	1	0.25	0.25
Cmax	mg/kg	11.9	11.3	69.6	101	15.7	18.7
Dose-norm  Cmax	mg/kg/mg*kg	0.595	0.565	0.347	0.505	0.785	0.935
AUClast	hr*mg/kg	70.0	53.3	843	680	101	90.8
Dose-norm AUClast	hr*kg*mg/kg/mg	3.50	2.67	4.22	3.40	5.05	4.54
AUC∞	hr*mg/kg	97.8*	59.9*	1345*	730*	229*	112*
Dose-norm  AUC∞	hr*kg*mg/kg/mg	4.89*	3.00*	6.73*	3.65*	11.5*	5.61*
% extrapolated	%	28.5	11.0	37.4	6.8	56.1	19.1
λz	1/hr	0.012	0.027*	0.008*	0.053*	0.006	0.020
t1/2	hr	60.5	26.1*	82.7*	13.2*	120	35.4
no. points		3	4	3	7	3	4
Corr coef.	r2	0.9615	0.8746	0.8535	0.6724	0.9795	0.9649

Conclusions:

No bioaccumulation potential based on study results. It can be concluded that Chloramine T administered orally was highly absorbed and excreted mainly via the urine. The data indicated that there were no sex differences in urinary, faecal or biliary excretion of radioactivity and no sex differences in total absorption. In addition, no differences were observed in urinary, faecal, biliary excretion or total absorption between the low and high dose or between single and repeated administrations. Linear plasma kinetics were observed. One major metabolite was observed in urine of all groups and identified as 4-Sulfamoyl benzoic acid. After oral administration of [methyl-14C]toluene-4 -sulphonamide (p-TSA) to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The 14C in the faeces was 4 -sulphamoylbenzoic acid, which probably originated in the tissues since the gut flora was unable to effect this biotransformation.The urine of rats given Chloramine–T contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%). These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Executive summary:

METHOD

Absorption, distribution, metabolism and excretion of [ring-U-14C]Chloramine T in the Wistar rat. The study was conducted according to the following guidelines: OECD Guideline no. 417: Toxicokinetics. Seven groups of rats were included in the study: Three (n=4 males and 4 females) for the massbalance, three (n=3 males and 3 females) for the toxicokinetics and one (n= 4 males and 4 females) for bile collection. Rats were dosed with a single oral dose of 20 mg/kg b.w. or 200 mg/kg b.w., or dosed repeatedly with 20 mg/kg b.w.for 10 days with unlabelled test substance

prior to a labeled dose. In the mass-balance groups, urine, faeces and bile (bile-cannulation group only) were collected in 0-8, 8-24, 24-48 and 48-72 hr intervals. Animals were euthanized 72 hours after dose administration, and several tissues and organs were collected. Total radioactivity in urine, faeces, bile, tissues and organs was determined. Selected urine and faeces samples were pooled per group and the metabolite profile in these pooled samples was investigated. In the toxicokinetic groups, blood was sampled from each rat at the following time points: 0.25, 0.5, 1 , 2, 4, 8, 24, 48 and 72 hour after dosing. Total radioactivity and Chloramine T equivalent concentrations were determined.

 

RESULTS

No mortality was observed in the study. One animal in the bile-cannulation group was killed in extremis, due to problems with the bile cannula. This animal displayed piloerection and hunched posture.

Clinical signs were observed mainly in the high dose ADME group and the bile cannulation group and consisted of piloerection.

From the plasma data it was evident that oral absorption proceeded fast, with Tmax values of 0.25 to 1 hour. Plasma concentration versus time curves of Chloramine T equivalents after oral dosing were similar, with similar Tmax values, and dose-normalised Cmax and AUC, in the same

order of magnitude. This suggests linear kinetics over the investigated dose range. Variable apparent terminal half-lives were observed, ranging from 13.2 to 120 hours. In general, it appeared that females had a shorter half-life of radioactivity than the males, but this was not reflected in the mass-balance data. The most important route of excretion of Chloramine T equivalents was urine. Urinary excretion accounted for 90% for males and 88% for females after low dosing, 81 % for males and 79% for females after high dosing and 87% for males and 78% for females after repeated dosing. In the bile-cannulation group urinary excretion accounted for 67% for the males and 66% for the females after oral dosing.

Faecal excretion was only a minor route of excretion. Faecal excretion accounted for 10% for the males and females after low dosing, 13% for the males and females after high dosing and 13% for the males and 16% for the females after repeated dosing, in the ADME groups. In the bile-cannulation group faecal excretion accounted for 25% for the males and 23% for the females. Biliary excretion accounted for 6% of the administered dose in males and 4% in females, in the bile-cannulation group. The absorption of Chloramine T equivalents was high, 92% and 91 % for the males and females in group 1 respectively, 83% in the males and females of group 2, and 89% and 85% of the males and females in group 3 respectively. The absorption in the bile-cannulation group was lower than in the ADME groups, 75% in both sexes. At termination of the study, the average total remaining radioactivity in blood, carcass plus tissues was between 0.6 and 1.4% of the administered dose in all groups..

The tissue concentration equivalents of Chloramine T were below the concentration observed in blood in all groups. 'This indicates that Chloramine T shows no potential for accumulation. The average total recovery of radioactivity in groups 1 to 3 and 7 was between 96 and 102% of

the administered dose. In the urine samples, one major radioactivity peak was observed, which accounted for 80-92% of the radioactivity in the radio-chromatogram and 65-81 % of the applied dose. This metabolite was isolated by prep HPLC from the urine pool of group 2 males and then identified as being 4. Sulfamoyl benzoic acid by NMR and Mass Spectrometry. Based on the radioactivity data (same major peak present in all groups and sexes at a similar retention time) it was concluded that the major radioactivity peak represented 4-Sulfamoyl benzoic acid in all groups.

In the radioactivity chromatograms of the faeces extracts 3 major peaks were observed, but these peaks could not be identified with MS. Two potential minor metabolites were identified i.e. p-sulfonic-benzoic acid and an aromatic methoxy compound.

 

CONCLUSIONS

It can be concluded that Chloramine T administered orally was highly absorbed and excreted mainly via the urine. The data indicated that there were no sex differences in urinary, faecal or biliary excretion of radioactivity and no sex differences in total absorption. In addition, no differences were observed in urinary, faecal, biliary excretion or total absorption between the low and high dose or between single and repeated administrations. Linear plasma kinetics were observed. One major metabolite was observed in urine of all groups and identified as 4-Sulfamoyl benzoic acid. After oral administration of [methyl-14C]toluene-4 -sulphonamide (p-TSA) to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The 14C in the faeces was 4 -sulphamoylbenzoic acid, which probably originated in the tissues since the gut flora was unable to effect this biotransformation.The urine of rats given Chloramine–T contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%). These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Reference 2

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

no data

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: The study was not performed according to a standard guideline and pre-GLP. Reporting is too limited to judge the validity of the study.

Objective of study:

metabolism

Principles of method if other than guideline:

No guideline followed.

GLP compliance:

no

Remarks:

pre-GLP

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: no data
- Age at study initiation: no data
- Weight at study initiation: 200-250g
- Fasting period before study: no
- Housing: Metabowls (Jencons Scientific Apparatus Limited, Hemd hempstead, Herts., U.K.) for the separate collection of urinc and faeces and allowed free access to food and water before and during the experinwnt.
- Individual metabolism cages: no data
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period:no data


ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): no data
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light):no data


IN-LIFE DATES: no data

Route of administration:

oral: gavage

Vehicle:

other: In 1.0 ml 20% ethanol in water or 50% aq. propylene glycol (1 ml)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
no data

VEHICLE
- Justification for use and choice of vehicle (if other than water): no data
- Concentration in vehicle: no data
- Amount of vehicle (if gavage): no data
- Lot/batch no. (if required): no data
- Purity: no data


HOMOGENEITY AND STABILITY OF TEST MATERIAL:
no data

Duration and frequency of treatment / exposure:

no data

Remarks:

Doses / Concentrations:
Females: 29 mg/kg 8uCi/rat; in 1.0 ml 20% ethanol in water, 200 mg/kg 15 uCi/rat; in 50% aq. propylene glycol (1ml)

No. of animals per sex per dose / concentration:

no data

Control animals:

no

Positive control reference chemical:

no

Details on study design:

no data

Details on dosing and sampling:

PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, expired air
- Time and frequency of sampling: no data

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sample: urine, faeces, expired air
- Time and frequency of sampling: no data
- From how many animals: samples were pooled, how many animals: no data
- Method type(s) for identification: Gas chromatography-mass spectrometry
- Limits of detection and quantification: no data

Statistics:

no data

After oral administration of [methyl-14C]toluene-4 -sulphonamide (p-TSA) to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The 14C in the faeces was 4 -sulphamoylbenzoic acid, which probably originated in the tissues since the gut flora was unable to effect this biotransformation.The urine of rats given Chloramine–T contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%).

Conclusions:

No bioaccumulation potential based on study results. After oral administration to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The urine of rats given p-TSA contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%). These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Executive summary:

After oral administration of [methyl-14C]toluene-4 -sulphonamide (p-TSA) to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The 14C in the faeces was 4 -sulphamoylbenzoic acid, which probably originated in the tissues since the gut flora was unable to effect this biotransformation.The urine of rats given Chloramine–T contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%). These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Reference 3

Endpoint:

dermal absorption in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

30 May 2006 - 01 September 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study performed accrding to guideline and under GLP.

Qualifier:

according to guideline

Guideline:

OECD Guideline 428 (Skin Absorption: In Vitro Method)

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Species:

human

Type of coverage:

open

Vehicle:

water

Duration of exposure:

8 hours

Doses:

300 or 50 µg/cm2, 3% or 0.5% solutions in water.

No. of animals per group:

6 skin membranes were used per dose and 2 skin membranes were prepared from each donor for each test group (A and B)

Control animals:

no

Details on in vitro test system (if applicable):

SKIN PREPARATION
- Source of skin: Human skin membranes were prepared from freshly excised skin obtained from three donors directly after abdominal surgery.
Donor 1: TNA 18/06, born in 1964, arrival at TNO on 26 June 2006
Donor 2: TNA 21/06, born in 1969, arrival at TNO on 28 August 2006
Donor 3: TNA 22/06, born in 1951, arrival at TNO on 30 August 2006
- Ethical approval if human skin: informed consent was provided by all donors.
- Type of skin: abdominal
- Preparative technique:
Human skin was dermatomed using a Dermatome 29 mm (Nouvag GmbH, Germany) to a recorded thickness of approximately 400μm. The exact thickness of all skin membranes was measured with a digimatic micrometer (Mirutoyo Corporation. Japan) and recorded.
- Thickness of skin (in mm): see: any other information on material and methods incl. tables
- Membrane integrity check: Not performed in order to start the experiment as soon as possible. An integrity test requires additional culturing time af the tissue wich may result in loss of viability.
- Storage conditions: The transportation of the skin to the laboratory was carried out as soon as possible after dissection (ca. 2 hours after receipt; skin from donor 1 was stored for about 4 hours in the refrigerator at the hospital until transport), while the skin was kept in a container placed on ice. After arrival at the laboratory, for all three donors, subcutaneous fat was removed and the skin dics were kept at 2-10 ºC during the night.
- Justification of species, anatomical site and preparative technique: The data is used for human risk assessment.

PRINCIPLES OF ASSAY
- Diffusion cell: The skin membranes were placed in 9 mm flow-through automated ditfusion cells (PenneGear Inc., Riegelsvilie, PA. USA).
- Receptor fluid: mixture of Dulbecco's Minimum Eaglc Medium (DMEM) and Ham F12 culture medium (3:l) supplemented with Epidermal Growth Factor (EGF, 10μg/L ), hydrocortisone (400 μg/L), gentamicin (50 mg/L) and Foetal Calf Serum (FCS, 10 %. v/v).
- Solubility of test substance in receptor fluid: The assumption was made that 10% of a 300 µg/cm2 ( = ca 192 µg a.i./skin membrane) dose reaches the recpetor fluid in 24 h. In this case the required solubility of Hlamid is 19.2 µg in 38.4 ml (flow rate ca 1.6 ml/h), i.e. approximately 0.5 µg/ml. The solubility of Halamid in water is 142 µg/ml and thus exceeds the required solubility by a factor 284. The solubilty of the degradation product p-TSA in water is 3 µg/ml and exceeds the required solubility by a factor 6. Therefore the solubility in the receptor fluid is considered to be sufficient.
- Static system: not applicable
- Flow-through system: The receptor fluid was pumped at a speed of cu 1.6 mL/h
- Test temperature: The mean skin surface tempenture was 32 ± 1 ºC
- Humidity: ambient humidity
- Occlusion: not applicable
- Reference substance(s): none

Absorption in different matrices:

see: remarks on results including tables and figures

Total recovery:

- Total recovery: The mean recovery was 95.4% (high dose) and 94.0% (low dose)
- Recovery of applied dose acceptable: yes
- Results adjusted for incomplete recovery of the applied dose: no
- Limit of detection (LOD):
- Quantification of values below LOD or LOQ:

Remarks on result:

other:

Stability:

In the receptor fluid, a rapid degradation of Chloramine-T trihydrate towards p-TSA was observed. A few minutes after preparing a 30mg/l solution 71.7% Chloramine-T trihydrate was left. The concentration further decreased from 48.9%, 29.2% to 14.5% after 7, 24 and 48 hours.

Directly after dosing, the dose solutions were analyzed by radio-HPLC. The relative amount of Chloramine-T trihydrate in the both dose solutions was 94 70 based on W detection. Based on radio-activity, this percentage was lower (ca  72%) due to additional peaks in the beginning of the radio-chromatogram  most probable related to a breakdown of the radio-label. Degradation will most likely take place to p-TSA (as is observed in the stability test using receptor fluid). It is not expected that only the radio-activity associated with the unknown peaks will be absorbed through the skin. A rapid a degradation of Chloramine-T trihydrate to p-TSA on the skin surface was confirmed by analyzing the cotton swab extracts. After 8 hours contact time, only ca 23 and ca 2.3 % Chloramine-T trihydrate was left in the coton swab extracts of the high and low dose group, respectively. It is therefore reasonable to assume that the main compound reaching the receptor fluid will be p-TSA. In the skin wash solution, Chloramine-T trihydrate was relatively stable deceted in time at 92% until 48 hours.

* The exact concetration is described in the method section.

**Total absorption is defined as the amount in the receptor fluid, the receptor compartment wash and the skin membrane, excluding the amount in the tape strips.

The in vitro percutaneous penetration of Halamid

Halamid	A		B
concentration measured (g/L)	30.0*		5.0*
Dose (µg/cm2)	300*		50 *
n	6		6
penetration into the receptor fluid after 24 h	% of dose	µg/cm2	% of dose	µg/cm2
	4.01	12.0	11.49	5.74
maximal flux (µg/cm2/h)	0.652		0.367
Lag time (h)	2.7		4.0
Total absorption (%)**	9.7		20.0

Conclusions:

The mean total adsorption of a 3% aqueous Chloramine-T trihydrate solution on human skin is 9.7%. The mean total adsorption of a 0.5% aqueous Chloramine-T trihydrate solution on human skin is 20.0%. These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Executive summary:

According to OECD 428 and under GLP the percutaneous absorption of  [14C]-Chloramine-T trihydrate in a 3% and 0.5% aqueous solution was evaluated on 6 freshly excised human membranes (2 from one donor). The chemical stability of Chloramine-T trihydrate was determined prior to the conduct of the study aiming at a later analysis of receptor fluid and skin wash fractions obtained in the main study. Skin membranes from three different human donors were used. The exposure time was 8 hours, after which the test compound was washed from the skin and the post-exposure time was 16 hours. Samples were taken from receptor fluid samples, skin wash. Receptor compartment wash, donor compartment wash, tape strips, and digested skin. All collected samples were analyzed with liquid scintillation counting (LSC).To determine the extent of (metabolic) degradation of Chloramine-T trihydrate into p-TSA, samples of the skin wash were analyzed by radio- LC. The total absorption is defined as the amount in the receptor fluid, the receptor compartment wash and the skin membrance, excluding the amount in the tape strips.

A rapid degradation of Chloramine-T trihydrate towards p-TSA was observed in the receptor fluid. Based on UV detection, the relative amount of Chloramine-T trihydrate decreased from 71.7 % established a few, minutes after mixing (t=0) to 14.5 % after 48 hours. In the skin wash solution, Chloramine-T trihydrate was relatively stable detected in time at 92% until 48 hours. For the high dose, the mean penetration of test compound-related radioactivity into the receptor fluid after 24 hours was 12.0 μg/cm2 which was 4.01% of the dose applied. The mean maximal flux was 0.652 μg/cm2/h  the lag time was 2.7 h. The mean total absorption, was 9.7%. For the low dose, the mean penetration of test compound-related radioactivity into the receptor fluid after 24 hours was 5.74 μg/cm2 which was 11.49 % of the dose applied The mean maximal flux was 0.367 μg/cm2/h and the lag time was 4.0 h. Slight differences between donors were observed. The mean total absorption was 20.0 %. The mean recovery was 95.4% (high dose) and 94.0 % (low dose). The mean total adsorption of a 3% aqueous Chloramine-T trihydrate solution on human skin  is 9.7%. The mean total adsorption of a 0.5% aqueous Chloramine T trihydrate solution on human skin is 20.0%. METHOD

Absorption, distribution, metabolism and excretion of [ring-U-14C]Chloramine T in the Wistar rat. The study was conducted according to the following guidelines: OECD Guideline no. 417: Toxicokinetics. Seven groups of rats were included in the study: Three (n=4 males and 4 females) for the massbalance, three (n=3 males and 3 females) for the toxicokinetics and one (n= 4 males and 4 females) for bile collection. Rats were dosed with a single oral dose of 20 mg/kg b.w. or 200 mg/kg b.w., or dosed repeatedly with 20 mg/kg b.w.for 10 days with unlabelled test substance

prior to a labeled dose. In the mass-balance groups, urine, faeces and bile (bile-cannulation group only) were collected in 0-8, 8-24, 24-48 and 48-72 hr intervals. Animals were euthanized 72 hours after dose administration, and several tissues and organs were collected. Total radioactivity in urine, faeces, bile, tissues and organs was determined. Selected urine and faeces samples were pooled per group and the metabolite profile in these pooled samples was investigated. In the toxicokinetic groups, blood was sampled from each rat at the following time points: 0.25, 0.5, 1 , 2, 4, 8, 24, 48 and 72 hour after dosing. Total radioactivity and Chloramine T equivalent concentrations were determined.

 

RESULTS

No mortality was observed in the study. One animal in the bile-cannulation group was killed in extremis, due to problems with the bile cannula. This animal displayed piloerection and hunched posture.

Clinical signs were observed mainly in the high dose ADME group and the bile cannulation group and consisted of piloerection.

From the plasma data it was evident that oral absorption proceeded fast, with Tmax values of 0.25 to 1 hour. Plasma concentration versus time curves of Chloramine T equivalents after oral dosing were similar, with similar Tmax values, and dose-normalised Cmax and AUC, in the same

order of magnitude. This suggests linear kinetics over the investigated dose range. Variable apparent terminal half-lives were observed, ranging from 13.2 to 120 hours. In general, it appeared that females had a shorter half-life of radioactivity than the males, but this was not reflected in the mass-balance data. The most important route of excretion of Chloramine T equivalents was urine. Urinary excretion accounted for 90% for males and 88% for females after low dosing, 81 % for males and 79% for females after high dosing and 87% for males and 78% for females after repeated dosing. In the bile-cannulation group urinary excretion accounted for 67% for the males and 66% for the females after oral dosing.

Faecal excretion was only a minor route of excretion. Faecal excretion accounted for 10% for the males and females after low dosing, 13% for the males and females after high dosing and 13% for the males and 16% for the females after repeated dosing, in the ADME groups. In the bile-cannulation group faecal excretion accounted for 25% for the males and 23% for the females. Biliary excretion accounted for 6% of the administered dose in males and 4% in females, in the bile-cannulation group. The absorption of Chloramine T equivalents was high, 92% and 91 % for the males and females in group 1 respectively, 83% in the males and females of group 2, and 89% and 85% of the males and females in group 3 respectively. The absorption in the bile-cannulation group was lower than in the ADME groups, 75% in both sexes. At termination of the study, the average total remaining radioactivity in blood, carcass plus tissues was between 0.6 and 1.4% of the administered dose in all groups..

The tissue concentration equivalents of Chloramine T were below the concentration observed in blood in all groups. 'This indicates that Chloramine T shows no potential for accumulation. The average total recovery of radioactivity in groups 1 to 3 and 7 was between 96 and 102% of

the administered dose. In the urine samples, one major radioactivity peak was observed, which accounted for 80-92% of the radioactivity in the radio-chromatogram and 65-81 % of the applied dose. This metabolite was isolated by prep HPLC from the urine pool of group 2 males and then identified as being 4. Sulfamoyl benzoic acid by NMR and Mass Spectrometry. Based on the radioactivity data (same major peak present in all groups and sexes at a similar retention time) it was concluded that the major radioactivity peak represented 4-Sulfamoyl benzoic acid in all groups.

In the radioactivity chromatograms of the faeces extracts 3 major peaks were observed, but these peaks could not be identified with MS. Two potential minor metabolites were identified i.e. p-sulfonic-benzoic acid and an aromatic methoxy compound.

 

CONCLUSIONS

It can be concluded that Chloramine T administered orally was highly absorbed and excreted mainly via the urine. The data indicated that there were no sex differences in urinary, faecal or biliary excretion of radioactivity and no sex differences in total absorption. In addition, no differences were observed in urinary, faecal, biliary excretion or total absorption between the low and high dose or between single and repeated administrations. Linear plasma kinetics were observed. One major metabolite was observed in urine of all groups and identified as 4-Sulfamoyl benzoic acid. After oral administration of [methyl-14C]toluene-4 -sulphonamide (p-TSA) to rats the label was rapidly eliminated largely in the urine (66 -89% dose) with little in the faeces (2 -8% dose). The 14C in the faeces was 4 -sulphamoylbenzoic acid, which probably originated in the tissues since the gut flora was unable to effect this biotransformation.The urine of rats given Chloramine–T contained 4– sulphamoylbenzoic acid as the major metabolite (93 % of the urinary 14C) together with small amounts of unchanged compound (1.5–2.3% of urinary 14C) 4–sulphamoylbenzyl alcohol (2.0–3.9%), 4 -sulphamoylbenzaldehyde (0–1.5 %) and at higher doses N–acetyltoluene–4–sulphonamide (2.1–2.3%). These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Reference 4

Endpoint:

dermal absorption in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Only a summary of the study.

Species:

other: Horse

A pharmacokinetic study in horses has been conducted to determine  concentrations of p-TSA after topical application 0.5 % Chloramine-T  solution. Six horses with wounds were treated twice a day at intervals of  8 hours over 72 h (6 treatments/per animal in total) by washing (3  animals, 200 ml to 1000 ml) or spraying (3 animals,100 to 250 ml). Blood  samples were collected immediately prior to each treatment, at 2, 4, 6,  and 8 h after the first treatment and at 2, 4, 6, 12, and 24 h after the  last treatment and analyzed with HPLC (LOD 3.2 µg/l) Chloramine-T was not  detected in any of the samples. p-TSA was only detectable in 2 samples  from the 5th treatment. Detected concentrations were 4.4 µg/l and 6.0  µg/l.

Conclusions:

Chloramine-T was not detected in any of the samples. p-TSA was only detectable in 2 samples from the 5th treatment. Detected concentrations were 4.4 µg/l and 6.0 µg/l. These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Executive summary:

A pharmacokinetic study in horses has been conducted to determine concentrations of p-TSA after topical application 0.5 % Chloramine-T solution. Six horses with wounds were treated twice a day at intervals of 8 hours over 72 h (6 treatments/per animal in total) by washing (3 animals, 200 ml to 1000 ml) or spraying (3 animals,100 to 250 ml). Blood samples were collected immediately prior to each treatment, at 2, 4, 6, and 8 h after the first treatment and at 2, 4, 6, 12, and 24 h after the last treatment and analyzed with HPLC (LOD 3.2 µg/l) Chloramine-T was not detected in any of the samples. p-TSA was only detectable in 2 samples from the 5th treatment. Detected concentrations were 4.4 µg/l and 6.0 µg/l. These results were used for read-across to N-ethyl-o (or p)-toluenesulfonamide.

Description of key information

Mostly by cross-reading from Chloramine-T:

It is shown that [14C]-Chloramine-T is almost instantaneously converted to p-TSA in contact with stomach and intestinal contents of rat. Similarly, also an in-vitro dermal absorption study demonstrated the conversion of Chloramine-T, resulting to only p-TSA to pass the skin.

Therefore the available ADME data on Chloramine-T is directly applicable in p-TSA.

Rapid and complete oral uptake followed by rapid excretion mainly with urine. Overall dermal absorption about 20%.

Key value for chemical safety assessment

Bioaccumulation potential:

no bioaccumulation potential

Absorption rate - oral (%):

100

Absorption rate - dermal (%):

20

Absorption rate - inhalation (%):

100

Additional information

Toxicokinetics

ADME data

Absorption, distribution, metabolism and excretion data onN-ethyl-o(or p)-toluenesulfonamideitself are not available and therefore the toxicokinetic assessment is based on the available toxicology andADME data from TSA isomers and N-chloro-4-methylbenzenesulfonamide sodium (Chloramine-T, CAS 127-65-1).

 

Information from physico chemical studies

An overview of the relevant physicochemical parameters for N-ethyl-o(or p)-toluenesulfonamide is provided in Physical and chemical properties

 

Absorption

The substance N-ethyl-o(or p)-toluenesulfonamide is in a neutral form at all physiological pH, and absorption is expected to be high.

 

Oral. Available ADME data on both p-TSA and o-TSA indicate rapid and complete oral uptake followed by rapid excretion mainly with urine, mainly as metabolite following oxidation of the methyl group on the benzene ring. In case of the ortho-position in o-TSA, the oxidized methyl and the amine can form a ring leading to the forming of saccharine, which is also found in the urine.

 

Dermal. Based on the water solubility and the low log Kow value of the substance N-ethyl-o(or p)-toluenesulfonamide high to moderate absorption could be expected (ECHA guidance, 7.12, Table R.7.12-3). Moreover, the small molecules with a molecular weight <500 are likely to be absorbed dermally. The dermal absorption is likely to be lower than the oral. For p-TSA, the estimated dermal penetration coefficient of 0.006 cm/h is comparable to that of water (0.0005 cm/hr). The estimated maximal dermal absorption is 0.0016 mg/cm²/hr (IH Skin Perm v1.03 from AIHA). For o-TSA the value for Maximal dermal absorption differs slightly with a lower value compared to p-TSA, which is related to lower water solubility

 

Cross-reading from results from in vitro dermal absorption from N-chloro-4-methylbenzenesulfonamide sodium (Chloramine-T), which basically measures the absorption of p-TSA, showed indicated that 20% absorption can be regarded as worst case. The level of absorption decreases with increasing concentrations, and at a concentration of 3% only 4% passed the skin and total absorption was about 10%.

 

Inhalation. There is no information on possible absorption via inhalation. However in view of low vapour pressure and low fraction of respirable particles in the products, respiratory exposures are not considered a great concern.Due to the very low vapour pressure any relevant uptake has to be by means of aerosols or particles. These will deposit mainly on upper airways, and will be subsequently swallowed following mucociliary transportation to pharynx. This results to no principal difference in absorption compared oral route.

 

Distribution

Based its molecular weightof the substance N-ethyl-o(or p)-toluenesulfonamideawider distribution is expected. As the low Log Kow value indicates hydrophilicity, the intracellular concentration is expected to be lower than the extracellular concentration (Particularly in fatty tissues). Following rapid absorption and excretion in urine, the concentrations of (metabolites of) the TSA substances is highest and longest present in the urinary tract, where therefore the toxic effects are first seen. Information available on benzenesulphonamides supports a common mechanism of action for toxicity to urinary tract observed in repeated dose studies with these compounds. The effect is indirect mediated by the appearance of calcium phosphate calculi caused by induced alkaline urine.

 

Metabolism

N-dealkylation and N-hydroxylation are the activated metabolic reactions for metabolic conversion of N-ethyl-o(or p)-toluenesulfonamide to TSA isomers. Under physiological conditions, an in vivo N-dealkylation process takes place through an N-oxidation intermediate step catalyzed by NADHP in the liver [7]. This reaction can produce TSA derivatives; o-TSA, p-TSA, and o/p-TSA by the N-dealkylation of N-ethyl-o(or p)-toluenesulfonamide. Alternatively, a non-enzymatic dealkylation may proceed catalytically in the presence of iron, producing o-TSA and p-TSA from N-ethyl-o(or p)-toluenesulfonamide [7]. This is supported by the In vivo Rat Metabolism Simulator available in QSAR Toolbox v.4.1 [8]. p-TSA and o-TSA are isomers, both structures are identical with the only difference, that is the position of the methyl-group on the benzene ring relative to the sulfonamide group. Toluenesulfonamide (TSA, methylbenzenesulfonamide) is the common substructure in all substances (Table 1).

 

Elimination

No substance specific information for N-ethyl-o(or p)-toluenesulfonamide regarding elimination is available. Furthermore, evaluation of systemic effects (acute toxicity and repeated dose toxicity) does not indicate any effects that could be linked to the route of excretion.

 

Accumulation

There is low potential to accumulate in individuals that are frequently exposed (e.g. daily at work) to the substance due to the low Log Kow values.

 

Conclusion

Oral uptake is expected based on information from the available test information (acute oral toxicity) and the range of physico chemical parameters. Dermal absorption would be possible based on physico-chemical information. Based its molecular weight, the distribution is expected. Fast absorption and subsequent elimination is confirmed by the available ADME data from TSA isomers and N-chloro-4-methylbenzenesulfonamide sodium (Chloramine-T, CAS 127-65-1). The absorption values for hazard assessment would be 100% for the inhalation route, 20% for the oral route and 100% for the dermal route, respectively.

Information is available from cross-reading from chloramine-T:

It is shown that [14C]-Chloramine-T is almost instantaneously converted to p-TSA in contact with stomach and intestinal contents of rat. Similarly, also an in-vitro dermal absorption study demonstrated the conversion of Chloramine-T, resulting to only p-TSA to pass the skin. Therefore the available ADME data on Chloramine-T is directly applicable in p-TSA.

 

ADME studies indicate that Chloramine-T (in the form of p-TSA) is rapidly and almost completely absorbed after oral administration, and relatively fast completely excreted again, mostly via the urine, as a benzoic acid derivative (4-sulfamoylbenozic acid, CAS 138-41-0). At termination, only about 1% of the activity remained in the body. About 78-87% is excreted in the urine, and 13-16% via the faeces. About 4-6% passes with the bile.

The tissue concentration of Chloramine-T equivalents (measured radiolabel) were below the concentration observed in blood in all groups indicating that there is no potential for accumulation. The availabe information from acute oral toxicity on p-TSA confirms this assessment. Clinical signs appear quick and at not too high dose levels resolve again within a day. At high dose levels pecifically involvent of the urinary tract is observed.

In-vitro dermal penetration studies with 0.5% aq Chloramine-T solution show that approximately 12% passed the skin completely and about 8% of the applied dose passed the stratum corneum but remained fixed in the skin after 24 hours. This results to an overall absorption of about 20%. Dermal absorption is thus lower then oral absorption. The amount of 20% can be regarded as worst case, as the level of absorption decreases with increasing concentrations: At a concentration of 3% only 4% passed the skin and total absorption was about 10%.

Metabolism: In the urine samples from Chloramine-T studies, one major radioactivity peak was observed. This peak was identified as benzoic acid derivative (4-sulfamoylbenozic acid, CAS 138-41-0). An older study from literature on p-TSA confirms the identity of the major metabolite (Ball, L.M., et al., 1978). Female albino Wistar rats were given a single low and high oral dose (29 mg/kg bw in 1ml 20% ethanol in water and 200 mg/kg bw in 50% aq. propylene glycol (1ml)). This study also identified the small amounts of other metabolites found next to a remaining small amount of parent compound p-TSA (1.5-2.3%). Other identified metabolites were 4-sulphamoylbenzyl alcohol (2.0-3.9%), 4-sulphamoylbenzaldehyde (0-1.5%) and at high dose N-acetyltoluene-4-sulphonamide (2.1-2.3%). Other studies reported in literature on metabolism were largely qualitative and confirm the metabolite4-sulfamoylbenozic acid in urine of dogs and rats (Flaschenträger B., et al., 1934; Minegishi, K. I., et al., 1972).

There is no information in uptake following inhalation. Due to the very low vapour pressure any relevant uptake has to be by means of aerosols or particles.These will deposit mainly on upper airways, and will be subsequently swallowed following mucociliary transportation to pharynx. This results to no principal difference in absorption compared oral route.