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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not specified
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference
Reference Type:
publication
Title:
Biocompatibility of EDTA, EGTA and citric acid
Author:
de Sousa, S. M. G. et al.
Year:
2005
Bibliographic source:
Braz. Dent. J. 16 (1): 3 - 8.

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Thirty-two adult male Wistar rats were used. Their backs were shaved and 2% Evans blue was injected intravenously into the lateral caudal tail vein. Immediately afterwards, 0.1 mL of citric acid solution was injected intradermally into the experimental site. The animals were assigned to 4 groups according to the time period tested. The animals were killed 30 minutes as well as 1, 3 and 6 hours after injection of the solution. The dorsal skin was dissected away and skin lesions were punched out. Each piece of skin containing the lesion was cut into small fragments and the dye was extracted upon immersion in 10 mL formamide. Then, the optical density was measured at 620 μm in a spectrophotometer The total amount of dye extracted from the samples was calculated by means of a standard calibration curve.
GLP compliance:
not specified
Remarks:
not specified in the publication

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
other: solution
Details on test material:
- State of aggregation: 15 % citric acid solution (supplier: Merck)
- pH: 1.0
Specific details on test material used for the study:
not specified

Test animals

Species:
rat
Strain:
Wistar
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: adult
- Weight at study initiation: approx. 350 g

Test system

Type of coverage:
open
Preparation of test site:
shaved
Vehicle:
other: deionized water
Controls:
yes
Amount / concentration applied:
not specified
Duration of treatment / exposure:
not applicable
Observation period:
30 minutes as well as 1, 3 and 6 hours after injection of the solutions
Number of animals:
32 male rats (8 animals/observation time point)
Details on study design:
NOTE: three further test item solutions were tested with the same animals as used for testing with the citric acid solution.

DOSAGE PREPARATION:
The tested acid solution was freshly prepared at the laboratory. The salt were weighed and diluted in deionized water, and pH was adjusted (pH meter B371, Micronal).

EXPERIMENTAL METHOD:
The rats were anesthetized with sulfuric ether, their backs were shaved. Their tails were washed and dried to facilitate the injections of 2% Evans blue (20 mg/kg) administered intravenously into the lateral caudal vein. Immediately afterwards, 0.1 mL of the test solution was injected intradermally into the experimental site. The animals were assigned to 4 groups according to the time period tested. The animals were killed 30 minutes, 1, 3 and 6 hours after injection of the solution. The dorsal skin was dissected away and skin lesions were punched out with a standard steel punch (3-cm diameter). Each piece of skin containing the lesion was cut into small fragments and the dye was extracted upon immersion in 10 mL formamide for 72 hours at 45 °C. After filtration with glass filter, the optical density was measured at 620 μm in a spectrophotometer (Ultrospec 2000, Pharmacia Biotech). The total amount of dye extracted from the samples was calculated by means of a standard calibration curve. Data were analyzed statistically by two-way ANOVA and Tukey’s HSD test.

Results and discussion

In vivo

Results
Remarks on result:
other: see "Remarks"
Remarks:
According to the authors, compared to saline (139.55 µg) a larger amount of dye was extracted after citric acid administration (329.81 µg), for all four periods evaluated. Furthermore, considering the time period, a significant difference (p<0.05) was observed between the 3 hour and 6 hour groups, but not between the 30-min and 1 hour groups. Also, the authors stated that there was an enhanced vascular permeability at the 6 hour experimental time. Although a statistically significant difference was seen between citric acid and saline, the means of dye extracted in these groups were numerically close to each other. Lastly, the authors stated that the results indicated that the citric acid solution yielded a continuous irritating potential even after 6 hours.
Irritant / corrosive response data:
Compared to saline (139.55 µg) a larger amount of dye was extracted after citric acid administration (329.81 µg), for all four periods evaluated.
Considering the time period, a significant difference (p<0.05) was observed between the 3 hour and 6 hour groups, but not between the 30-min and 1 hour groups. There was an enhanced vascular permeability at the 6 hour experimental time.
Although a statistically significant difference was seen between citric acid and saline, the means of dye extracted in these groups were numerically close to each other.
The results indicated that the citric acid solution yielded a continuous irritating potential even after 6 hours.

Please aslo refer to the field "Any other information on results incl. tables" below.

Any other information on results incl. tables

Table 1: Means (μg) and standard deviations (± SD) of the total amount of dye extracted in the control and experimental groups.

Time

Citric acid

Saline

30 minutes

280.92 ± 140.84

153.93 ± 47.15

1 hour

271.41 ± 135.09

115.68 ± 48.57

3 hours

286.43 ± 102.08

115.36 ± 58.60

6 hours

480.50 ± 212.34

173.23 ± 51.31

Applicant's summary and conclusion

Conclusions:
According to the authors, compared to saline (139.55 µg) a larger amount of dye was extracted after citric acid administration (329.81 µg), for all four periods evaluated. Furthermore, considering the time period, a significant difference (p<0.05) was observed between the 3 hour and 6 hour groups, but not between the 30-min and 1 hour groups. Also, the authors stated that there was an enhanced vascular permeability at the 6 hour experimental time. Although a statistically significant difference was seen between citric acid and saline, the means of dye extracted in these groups were numerically close to each other. Lastly, the authors stated that the results indicated that the citric acid solution yielded a continuous irritating potential even after 6 hours.