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Exposure related observations in humans: other data

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Administrative data

Endpoint:
exposure-related observations in humans: other data
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Publication, not fully documented; iron salt was used in watery solution exceeding by far the water solubility so that flocculation and mechanical hindrance are likely effect triggers

Data source

Reference
Reference Type:
publication
Title:
In vitro effects of metal ions (Fe2+, Mn2+, Pb2+) on sperm motility and lipid peroxidation in human semen
Author:
Huang YL, Tseng WC, Lin TH
Year:
2001
Bibliographic source:
DOI 10.1080/009841001459414 PMID 11245395 J Tox Env Health 62(4):259-67

Materials and methods

Type of study / information:
In vitro effect of iron salt solutions on human sperm motility and lipid peroxidation
Endpoint addressed:
toxicity to reproduction / fertility
Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
In vitro study with human semen to investigate the effects of metal ions on sperm motility and lipid peroxidation. Human sperm was incubated with ferrous nitrate at concentrations of 5, 50 and 500 ppm.
Sperm motility was determined by microscopy, and lipid peroxidation was assessed by determination of MDA by HPLC.
MDA (malondialdehyde) is used as marker of lipid peroxidation.
In the publication apart of ferrous nitrate, also effects of lead nitrate and manganese nitrate was investigated but not reported in the RSS.
GLP compliance:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): ferrous nitrate Fe(NO3)2 x 9H2O purchased from Merck, Germany

Method

Ethical approval:
not applicable
Details on study design:
Fresh semen samples provided from 5 healthy volunteers were evaluated according to the WHO Guidelines (1992) including volume, sperm concentration, percent motility, and sperm morphology. Semen samples with leukocyte-to-spermatozoa concentration of <5 % were used for the test. All samples were collected by masturbation after 3 to 4 days of abstinence.
135 µL semen was mixed with 15 µL of saline (control) or 50, 500 or 5000 ppm of metal ion solution.
For the time-course experiment: semen samples were incubated for 2, 4, 6, or 8 hours with saline or with metal ions. For the concentration-effect experiment: semen samples were incubated for 2 to 8 hours with saline or with 5, 50 or 500 ppm metal ions. For the study on MDA levels: Semen were treated with metal ions for 8 h. After 8 h of treatment, the semen was centrifuged at 7000 x g for 15 min to separate the sperm and sperm plasma.
Sperm Motility Assay:
Motility was defined as percentage of sperm count with light microscopy. At least 200 spermatozoa per sample were examined. All motile sperm were viable and considered vigorous.
Lipid Peroxidation Assay:
Duplicate aliquots of sperm plasma (50 µL) were analysed for MDA by thiobarbituric acid reaction. With HPLC separation of the MDA-TBA adduct using tetra-ethoxypropane as standard, according to Wong et al. (1987).
Statistical evaluation
All values presented as mean SD. Statistical significance was evaluated using Student’s t-test when control and metal ion-treated semen were compared for the sperm motility and lipid peroxidation after2 to 8 h incubation. Level of significance at p < 0.05
Exposure assessment:
estimated
Details on exposure:
EXPOSURE LEVELS: ferrous nitrate at concentrations of 5, 50 and 500 ppm
EXPOSURE PERIOD:
-Time-course experiment: semen samples were incubated for 2, 4, 6, or 8 hours with control (saline) or with metal ion
-Concentration-effect experiment: semen samples were incubated for 2 to 8 hours with control (saline) or with metal ion
-Study on MDA levels: Semen were treated with metal ions for 8h
POSTEXPOSURE PERIOD:
None, evaluation was made immediately

Results and discussion

Results:
Reduction in sperm motility was associated with lipid peroxidation: significant inhibition of motility was observed at 5 ppm Fe2+ associated with a marked rise in malondialdehyde (MDA).

Any other information on results incl. tables

Exposure to 5 ppm Fe2+for 6 h resulted in a decrease of sperm motility. Exposure to 50 and 500 ppm Fe2+significantly reduced sperm motility at 4 and 2 h, respectively which maintained for 8 h. Fe2+significantly increased MDA levels in seminal plasma starting at the low dose.

The iron salt was used in watery solution exceeding by far the water solubility so that flocculation and mechanical hindrance are likely effect triggers

Applicant's summary and conclusion

Conclusions:
Reduction in sperm motility was associated with lipid peroxidation: significant inhibition of motility was observed at 5 ppm Fe2+ onwards associated with a marked rise in malondialdehyde (MDA).
Executive summary:

An in vitro study with human semen was conducted with ferrous nitrate (Fe(NO3)2x 9H2O) in order to investigate the effects of Fe2 +ions on sperm motility and lipid peroxidation. Human sperm was incubated with ferrous nitrate at concentrations of 5, 50 and 500 ppm. Sperm motility was determined by microscopy, and lipid peroxidation was assessed by determination of MDA by HPLC. MDA (malondialdehyde) is used as marker of lipid peroxidation.

Exposure to 5 ppm Fe2+for 6h resulted in a decrease of sperm motility. Exposure to 50 and 500 ppm Fe2+significantly reduced sperm motility at 4 and 2h, respectively which maintained for 8h. Fe2+significantly increased MDA levels in seminal plasma starting at the low dose. However, it is concluded by the applicant that this study is only of minor relevance for living organisms.

The iron salt was used in watery solution exceeding by far the water solubility so that flocculation and mechanical hindrance are likely effect triggers