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Toxicological information

Specific investigations: other studies

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Administrative data

Endpoint:
hematoxicity
Type of information:
experimental study
Adequacy of study:
other information
Study period:
No data
Reliability:
2 (reliable with restrictions)
Cross-referenceopen allclose all
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study

Data source

Reference
Reference Type:
publication
Title:
Effect of exogenous glutathione, glutathione reductase, chlorine dioxide and chlorite on osmotic fragility of rat blood in vitro.
Author:
Abdel-Rahman, M.S.; Couri, D. and Bull, R.J.
Year:
1984
Bibliographic source:
Journal of the American College of Toxicology, 3(4):269-275

Materials and methods

Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Rats were sacrificed by decapitation and blood was collected in heparinised tubes. Vessels containing 3 mL of blood each were incubated in water bath at 37°C after the addition of ClO2. The effect of exogenous glutathione, glutathione reductase, chlorine dioxide and chlorite on osmotic fragility of rat blood were investigated in vitro.
Osmotic fragility determination: The addition of blood to hypotonic solution was used to determine the osmotic fragility by measuring the concentration of hemoglobin released in the solution by the method of Dacie (1977).
Reagent: Stock solution of buffered sodium chloride was prepared by dissolving 90 g of NaCl, 13.66 g of Na2HPO4, and 2.43 g of NaH2PO4 in 1 L double-distilled water. This stock solution is osmotically equivalent to 10% NaCl. The working solution was prepared by dilution of the stock solution 1:10 to make up a solution equivalent to 1%
Assay procedures: 5 mL of the various working solutions was added to the test tubes. 50 µL of blood was added to each tube and mixed well. After 30 minutes, tubes were centrifuged at 500 g for 5 minutes. The osmotic fragility curve in rat blood was plotted as percent hemolysis against salt concentration.
Effect of exogenous glutathione and ClO2 on osmotic fragility of rat blood: Reduced glutathione (50 mg%) was added to rat blood directly before the addition of ClO2. The final concentration of ClO2 was 100 mg/L. Osmotic fragility was determined after 1 and 2 hours at 37°C as described above.
Effect of glutathione reductase and ClO2 on osmotic fragility of rat blood: ClO2 (100 mg/L) was added to rat blood, afterwards 17 units (170 µg) of glutathione reductase and 20 µL NADPH (2mM) were added. Tubes were incubated at 37°C and 50 µL aliquots withdrawn at 1 hour to determine osmotic fragility.
GLP compliance:
not specified
Type of method:
in vitro

Test material

Constituent 1
Chemical structure
Reference substance name:
Chlorine dioxide
EC Number:
233-162-8
EC Name:
Chlorine dioxide
Cas Number:
10049-04-4
Molecular formula:
ClO2
IUPAC Name:
Chlorine Dioxide
Details on test material:
No data

Test animals

Species:
rat
Strain:
not specified
Sex:
not specified
Details on test animals or test system and environmental conditions:
No data

Administration / exposure

Details on exposure:
No data
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
No data
Duration of treatment / exposure:
No data
Frequency of treatment:
No data
Post exposure period:
No data
No. of animals per sex per dose:
No data
Details on study design:
No data

Examinations

Examinations:
No data
Positive control:
No data

Results and discussion

Details on results:
RBC hemolysis was decreased in rat blood after 30, 60 and 120 minutes. The glutathione content expressed as percentage of controls was decreased with incubation time. When ClO2 was added with reduced glutathione (GSH) to the blood, no effect on hemolysis was observed with ClO2 treatment alone. Addition of NADPH alone prevented ClO2 from exhibiting hemolysis resistance, while glutathione reductase (GR) and its cofactor (NADPH) increased hemolysis about 1.5-2 fold. Removing GR only resulted in increased resistance to hemolysis with ClO2. The formation of disulfide bonds between sulfhydryl groups in erythrocytic membranes and hemoglobin, causing precipitation of hemoglobin (yielding apparent resistance to hemolysis) can account for the difference between the hemolysis before and after the addition of GR.

Any other information on results incl. tables

No data

Applicant's summary and conclusion

Executive summary:

In an in vitro study, rat blood cells were exposed to ClO2 alone or combined with exogenous glutathione or glutathione reductase by adding 5 mL of the various working solutions to 50 µL of collected blood.

The osmotic fragility and glutathione content was studied in rat blood treated with CIO2 in vitro. RBC hemolysis was decreased in rat blood after 30,60, and 120 minutes by all treatments. The glutathione content expressed as percentage of controls was decreased with incubation time. When CIO2 was added with reduced glutathione (GSH) to the blood, no effect on hemolysis was observed compared to control or to GSH alone at 2 hours, but decreased hemolysis was observed with CIO2 treatment alone. Addition of NADPH alone prevented CIO2 from exhibiting hemolysis resistance, while glutathione reductase (GR) and its cofactor (NADPH) increased hemolysis about 1.5–2 fold. Removing GR only resulted in increased resistance to hemolysis. The formation of disulfide bonds between sulfhydryl groups in erythrocyticmembranes and hemoglobin, causing precipitation of hemoglobin (yielding apparent resistance to hemolysis) can account for the difference between the hemolysis before and after the addition of GR.