Chlorine dioxide

Specific investigations: other studies

Administrative data

Endpoint:

hematoxicity

Type of information:

experimental study

Adequacy of study:

other information

Study period:

No data

Reliability:

2 (reliable with restrictions)

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

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

Test material information

Constituent 1

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.