4-ethylphenol

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

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

Adequacy of study:

other information

Study period:

2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

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

Justification for type of information:

Justification for read-across attached

Data source

Reference

Reference Type:

publication

Title:

Quantitative analysis of cresol and its metabolites in biological materials and distribution in rats after oral administration

Author:

Yasumasa Morinaga et al.

Year:

2004

Bibliographic source:

Legal Medicine 6 (2004) 32–40 (www.elsevier.com)

Materials and methods

Objective of study:

distribution

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

No guideline stated in the publication. Oral application of a cresol soap solution to male rats via gastric tube and examination of cresol content in various tissues at various time points as well as the analysis of the respective glucuronides and sulfates.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

p-cresol

EC Number:

203-398-6

EC Name:

p-cresol

Cas Number:

106-44-5

Molecular formula:

C7H8O

IUPAC Name:

p-cresol

Constituent 2

Reference substance name:

m-cresol

EC Number:

203-577-9

EC Name:

m-cresol

Cas Number:

108-39-4

Molecular formula:

C7H8O

IUPAC Name:

m-cresol

Test material form:

other: Diluted cresol soap solution

Details on test material:

cresol soap solution containing 200 mg/mL p-cresol and 320 mg/mL m-cresol

Radiolabelling:

not specified

Test animals

Species:

rat

Strain:

Wistar

Sex:

male

Details on test animals or test system and environmental conditions:

Male Wistar rats (average weight 410g) fed on standard diet with free access to tap water were fasted for 24 h before administration.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Duration and frequency of treatment / exposure:

single treatment

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

100 mg/kg bw (total dose)

Remarks:

p-cresol

Doses / concentrations 2

Dose / conc.:

160 mg/kg bw (total dose)

Remarks:

m-cresol

No. of animals per sex per dose / concentration:

Not stated in the publication.

Control animals:

not specified

Positive control reference chemical:

Not stated in the publication.

Details on study design:

A cresol soap solution (p-cresol 200 mg/ml, m-cresol 320 mg/ml containment) was bought at a drugstore and it was diluted with water to one-fifth concentration for use.
The rats were dosed with the diluted cresol soap solution, of which amounts were 2.5 ml/kg (100 mg p-cresol and 160 mg m-cresol per kg), via gastric tube.
At various times after administration, blood, brain, lung, liver, spleen, kidney, femoral muscle and gastric contents were obtained immediately after sacrifice by cervical dislocation. The specimens were frozen at -30°C as quickly as possible, and preserved until analysis.
The quantitative analysis method for the measurement of the concentration of cresol, cresol-glucuronide and cresol-sulfate in organs was investigated using the rat organs obtained at 4 h after administration and the urine collected from a cresol poisoning patient.

Cresol analysis conditions
The high-performance liquid chromatograph (HPLC) system used was composed of a pump (LC-10AS, Shimadzu Co. Ltd., Kyoto, Japan), a fluorescence detector (RF-10AXL, Shimadzu Co. Ltd.) and an auto-sample injector (AS-100, Bio-Rad, CA).

Statistics:

Not stated in the publication.

Results and discussion

Preliminary studies:

Not applicable

Toxicokinetic / pharmacokinetic studies

Details on absorption:

Not applicable

Details on distribution in tissues:

cresol administered by gavage to male Wistar rats were detected in blood and distributed to the brain, kidney, liver, lung, muscle, and spleen.

Details on excretion:

The unconjugated cresol as well as the glucuronide and sulfate metabolites were eliminated from blood, brain, lung, muscle, liver, spleen, and kidneys for the most part within several hours.

Metabolite characterisation studies

Metabolites identified:

yes

Details on metabolites:

m-cresol was easily metabolized to cresol sulfate, and the p-cresol to cresol glucuronide.
The metabolism of cresol is obviously dependent on its isomers. That is why the time course changes of cresol metabolites in blood vary between p-cresol and m-cresol.

Any other information on results incl. tables

Confirmation of endogenous hydrolysis enzyme of cresol conjugates in tissues

m-Cresol concentration in the liver homogenate increased gradually over 24 h. The concentration in the liver homogenate with b-glucuronidase went up sharply for 2 h and then kept constant over 24 h. The concentration in the liver homogenate with sulfatase went up sharply for 1 h and then increased gradually over 24 h. After incubation for 24 h, the concentration in the liver homogenate with b-glucuronidase and that in the liver homogenate were much the same. The concentration in the homogenate with sulfatase was higher than that in the liver homogenate with a constant difference.

The homogenates of brain, lung, spleen, kidney and femoral muscle of the rat which had been fed cresol were adjusted to pH 5.0 or 7.5 and incubated at 37°C for 2 h. m-Cresol concentrations were compared to that in the homogenate without incubation. m-Cresol concentrations in the lung and the kidney homogenates at pH 5.0 increased during the incubation for 2 h. At pH 7.5, m-cresol concentration did not increase in any of the homogenates.

Effect of saccharolactone on cresol-sulfate analysis

The effect of saccharolactone (0.2, 0.4 and 1.0 mmol) to 500 units of b-glucuronidase was investigated using diluted urine collected from a cresol poisoned patient. Even 0.2 mmol of saccharolactone inhibited the b-glucuronidase activity completely.

The effect of saccharolactone on liver homogenate was investigated using each liver sample collected from five rats which had been fed cresol. The homogenates adjusted at pH 7.5 were incubated with or without saccharolactone (0.4 mmol) at 37°C for 2 h. Cresol concentration in the homogenate without saccharolactone increased, while that in

the homogenate with saccharolactone did not increase at all .

m-Cresol concentration increased during the first hour and then remained constant for 4 h.

Accuracy and precision of this method

The linear calibration curves were obtained in the concentration range of 0.5–100 mg/ml for p-cresol and m-cresol. The correlation coefficient of the curve was 0.999 for both cresols. The minimum detection level in aqueous solution was 0.1 mg/mL in both p-cresol and m-cresol.

The concentration of each compound was calculated from the calibration curve. The coefficients of variation of 1.0 mg/g with various treatments ranged from 2.3 to 12.2%, respectively, and those of 10 and 100 mg/g were within 2.5% in each treatment.

The errors of the present method were within 10% of the spiked value.

The residual rate of the dosed cresol in gastric contents

The residual rate is a percentage of the cresol quantity recovered from the gastric contents to the dose amount of cresol. Approximately 50% of the administered cresol disappeared 15 min after administration, and then gradually disappeared from the stomach. Almost all administered cresol disappeared within 8 h.

Time course changes of the cresol and its metabolites in the blood

The unconjugate concentrations of p-cresol and m-cresol decreased rapidly for 2 h after peaking at 30 min after administration. The unconjugates were not detected after 4 h. The p-cresol concentrations of glucuronide were always higher than those of sulfate.

The m-cresol concentrations of sulfate were consistently higher than the glucuronide except at the 1 h point.

The concentration ratio of m-cresol to p-cresol in the cresol soap solution used in this experiment was 1.6:1. The ratio of the unconjugate in blood was 2.2 (minimum–maximum: 1.8–3.5) on the average, glucuronide was 0.8 (0.5–1.2), and sulfate was 3.6 (1.2–6.2).

Time course changes of the cresol and its metabolites in tissues

The unconjugate concentrations of both cresols in the brain, lung and muscle were approximately equal to those in blood, and the time course changes of each accorded with that of the blood. The concentration ratio of the blood to the brain was 1.3 (0.7–2.0), to the lung was 1.1 (0.6–2.2), and to the muscle was 0.8 (0.5–2.2). In the liver and spleen the concentrations of unconjugate were always much higher than those in blood.

The p-cresol and m-cresol concentrations of glucuronide were always the highest in the kidney followed by the liver. The concentrations of glucuronide in the lung and blood were nearly equal at any given time. The glucuronide concentrations in the brain and muscle were lower in comparison to the other organs, especially the brain.

The p-cresol and m-cresol concentrations of sulfate in the brain, lung and muscle were lower than those in blood. In the brain and muscle, the sulfate concentrations were only about 10–20% of those in the blood. In the lung, they were about 60% of those in the blood, and the time course change of the lung coincided with that of blood. The sulfate concentration in the kidney was highest at 15 min, thereafter fell off rapidly for 1 h, and was almost regular in concentration after 1 h. The p-cresol sulfate concentrations in the kidney were nearly equal to those in blood. The m-cresol sulfate concentrations in the kidney were always the highest of all tissues; the values were about two times higher than blood levels.

In a comparison between the concentrations of glucuronide in tissues and those of sulfate, the glucuronide was almost always higher than the sulfate in both p-cresol and m-cresol, especially in the liver and kidney. p-Cresol glucuronide was, on average, 12 times (1–36) higher than sulfate in liver, and 15 times (2–76) higher than in kidney.

m-Cresol of glucuronide was on average nine times (2–35) higher than sulfate in liver, and two times (1–5) higher in kidney.

Applicant's summary and conclusion

Conclusions:

The unconjugates of cresol in the liver, spleen and kidney were detected in high concentrations even when the unconjugates were not detected in the blood.
m-Cresol was easily metabolized to sulfate, and the p-cresol to glucuronide in rats. The concentration ratio of m-cresol to p-cresol in blood and organs was different from the rate of the cresol soap solution that was administered. The pharmacokinetics was different between p-cresol and m-cresol in rats.