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EC number: 941-679-1 | CAS number: -
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Basic toxicokinetics
Administrative data
- Endpoint:
- basic toxicokinetics in vivo
- Type of information:
- experimental study
- Adequacy of study:
- supporting study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: well-documented publication, which meets basic scientific principles
Data source
Reference
- Reference Type:
- publication
- Title:
- Chelation of Mitochondrial Iron Prevents Seizure-Induced Mitochondrial Dysfunction and Neuronal Injury
- Author:
- Liang, L., Jarrett, S.G. and Patel M.
- Year:
- 2 008
- Bibliographic source:
- The Journal of Neuroscience, November 5, 2008 - 28(45):11550 –11556
Materials and methods
- Objective of study:
- distribution
Test guideline
- Qualifier:
- no guideline followed
- Principles of method if other than guideline:
- Adult male Sprague Dawley rats were injected subcutaneously (s.c.) with a single dose (12 mg/kg) of kainic acid (KA) dissolved in sterilized saline. HBED was injected at the dose of 75 µmol/kg, s.c. daily three times before KA and once after KA for a total of four injections. Thereafter the total brain iron levels were assayed.
- GLP compliance:
- not specified
Test material
- Reference substance name:
- N,N-bis (2-hydroxybenzyl) ethylenediamine-N,N-diacetic acid
- IUPAC Name:
- N,N-bis (2-hydroxybenzyl) ethylenediamine-N,N-diacetic acid
- Details on test material:
- - Name of test material (as cited in study report): N,N-bis (2-hydroxybenzyl) ethylenediamine-N,N-diacetic acid (HBED)
- Molecular formula (if other than submission substance): C20H24N2O6
- Molecular weight (if other than submission substance): 424.89 g/mol
- Smiles notation (if other than submission substance): OC(=O)CN(CCN(CC(=O)O)Cc1ccccc1O)Cc1ccccc1O
- Substance type: chelate
- Physical state: solid
Constituent 1
- Radiolabelling:
- no
Test animals
- Species:
- rat
- Strain:
- Sprague-Dawley
- Sex:
- male
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Age at study initiation: 3 month of age
- Weight at study initiation: 300–350 g
Animal studies were performed in accordance with the National Institute of Health Guide for the Care and Use of Laboratory Animals (NIH Publications No. 80-23). All procedures were approved by the Institute Animal Care and Use Committee (IACUC) of the University of Colorado at Denver and Health Sciences Center (UCDHSC), which is fully accredited by the American Association for the Accreditation of Laboratory Animal Care.
Administration / exposure
- Route of administration:
- subcutaneous
- Vehicle:
- other: dissolved in dimethyl sulfoxide (DMSO) and diluted with sterilized phosphate buffered saline (PBS) to achieve the desired final concentration (1% DMSO)
- Details on exposure:
- Kainic acid (KA) and N,N-bis (2-hydroxybenzyl) ethylenediamine-N,N-diacetic acid (HBED) administration
Adult male Sprague Dawley rats (3 months of age, weighing 300–350 g) were injected subcutaneously (s.c.) with a single dose (12 mg/kg) of KA (Ocean Products International) dissolved in sterilized saline. HBED (Strem Chemicals) was injected at the dose of 75 µmol/kg, s.c. daily three times before KA and once after KA for a total of four injections. HBED was dissolved in dimethyl sulfoxide (DMSO) and diluted with sterilized phosphate buffered saline (PBS) to achieve the desired final concentration (1% DMSO). - Duration and frequency of treatment / exposure:
- HBED (Strem Chemicals) was injected at the dose of 75 µmol/kg, s.c. daily three times before KA and once after KA for a total of four injections.
Doses / concentrations
- Remarks:
- Doses / Concentrations:
0 mol/kg, 75 µmol/kg
- No. of animals per sex per dose / concentration:
- 3-6 animals
- Control animals:
- yes, concurrent vehicle
- Details on dosing and sampling:
- PHARMACOKINETIC STUDY (distribution)
- Tissues and body fluids sampled: brain tissue
- Time and frequency of sampling: rats were killed 7 d after KA administration
Results and discussion
Main ADME results
- Type:
- distribution
- Results:
- Intravascular compartment
Toxicokinetic / pharmacokinetic studies
- Details on absorption:
- No detailed information could be obtained in this study as the chemical was administered subcutaneously.
- Details on distribution in tissues:
- HBED penetrates the BBB and mitochondria - Before evaluating its neuroprotective effects, they determined the brain bioavailability of HBED after systemic administration. They measured HBED concentrations in the forebrain and mitochondrial fractions isolated from the hippocampi of rats at different time points after a single injection (75 µmol/kg, s.c). HBED levels were detected 1 h after administration, peaked at 3 h and remained stable for at least 6 h. Both tissue and mitochondrial levels remained 25% of initial values at the 24 h time-point after injection. These results suggest that HBED penetrates the BBB and is accessible to hippocampal mitochondria. The total iron levels in the rat hippocampus were measured with varying doses of HBED. Total iron levels measured by the ferrozine method in the hippocampus were decreased 30% after 4 or 5 daily injections with 75 µmol/kg. Increasing the injection frequency to 7 resulted in a 38% reduction of iron content compared with control animals but retarded weight gain. The dose of HBED of 75 µmol/kg was further optimized based on a dose–response study using chelatable iron changes in the mitochondria as an endpoint.
Transfer into organs
- Transfer type:
- blood/brain barrier
- Observation:
- distinct transfer
Metabolite characterisation studies
- Metabolites identified:
- no
Any other information on results incl. tables
HBED administration does not alter KA-induced behavioural seizure indices
To determine whether the HBED treatment protocol influenced KA-induced behavioural seizures, they evaluated seizure scores, latency to wet dog shakes and duration of SE. No significant differences were observed in behavioural seizure intensity, seizure latency time (44.0 +/-3.4 and 45.3 +/- 3.2 min for KA and KA-HBED, respectively; n = 12 rats per group) and seizure duration (384.2+/- 15.4 and 388.3+/- 16.1 min for KA and KA-HBED, respectively; n= 12 rats per group). The percentage of animals that received KA or KA-HBED treatment that did not go on to exhibit a stage 3 or 4 seizure and were thus eliminated from the study was 3.4% (2 of 59 rats) and 3.3% (2 of 60 rats), respectively. Mortality from KA alone and KA-HBED was 5.1% (3 of 59 rats) and 6.7% (4 of 60 rats), respectively. No significant differences in mortality rate were observed between KA and KAHBED groups.
HBED inhibits seizure-induced chelatable mitochondrial iron changes
Next, they assessed the effect of different doses of HBED on KA-induced increases in mitochondrial chelatable iron 24 h after KA injection in the four treatment groups. HBED doses of 75 and 150 µmol/kg, but not 37.5 µmol/kg, for a total 4 injections significantly decreased KA-induced changes in chelatable iron in the hippocampal mitochondrial fractions. Based on the bioavailability parameters and the ability to decrease total as well as chelatable iron, subsequent studies were conducted with the daily HBED dose of 75 µmol/kg for 4 d. The effects of HBED on SE-induced iron chelation were further verified using the RPA fluorescence method. RPA fluorescence quenching has been shown to be a selective indicator of intramitochondrial chelatable iron in both tissue and cells. HBED treatment significantly attenuated the KA-induced increase in mitochondrial chelatable iron detected by RPA fluorescence quenching. Analysis of RPA fluorescence density revealed that HBED treatment most prominently attenuated the increase in chelatable iron in the hippocampal CA3 region but also in the CA1 and dentate hilus.
HBED protects against SE-induced mitochondrial dysfunction
To determine whether mitochondrial iron chelation by HBED was sufficient to preserve mitochondrial targets after KA treatment, they analysed the integrity of mtDNA using a sensitive QPCR assay specific to the mitochondrial genome. KA-induced SE generated an approximate fivefold increase in mtDNA damage in the hippocampus compared with the control group at the 24 h time point. SE-induced mtDNA damage was significantly attenuated by HBED.
HBED inhibits SE-induced mitochondrial oxidative stress
To further determine the deleterious role of mitochondrial iron in SE-induced injury, they measured two additional indices of mitochondrial oxidative stress, the GSH, GSSG and 8-OHdG/2dG ratios. Twenty four hours after KA injection, hippocampal mitochondrial GSH levels were depleted ca. 40%; whereas the corresponding levels of GSSG were significant increased ca. 240% compared with controls. The changes in both GSH and GSSG were significantly restored by HBED. The level of DNA oxidation measured by the ratios of 8-OHdG/2dG, was enhanced > 3-fold by KA-induced SE and attenuated ca. 50% by HBED in the hippocampal mitochondrial fractions.
HBED protects against SE-induced neuronal degeneration
Finally, they evaluated the neuroprotective effects of HBED by Fluoro-Jade B fluorescence, a sensitive marker assessing degeneration of neuronal cell bodies, dendrites, axons, or terminals. Previous reports in the literature have demonstrated that Fluoro-Jade B is a more sensitive, reliable and definitive marker of neuronal degeneration than silver staining techniques. No significant Fluoro-Jade B staining indicative of degeneration was observed in any brain region of control animals. However, significant staining (degeneration) was observed in the cell bodies and terminals in the hippocampal CA1, CA3 and hilar regions but not granule cell layer of rats injected with KA beginning ca. 48 h and peaking at ca. 7 d after injection. The percentage of relative fluorescence density quantified by Image J was increased ca. 190, ca. 240 and ca. 265% in the CA1, CA3 and hilus, respectively in the KA vs control groups. Significant protection of neuronal degeneration was observed in the KAHBED group compared with KA alone indicating a neuroprotective effect of HBED.
Applicant's summary and conclusion
- Conclusions:
- Interpretation of results (migrated information): other: Intravascular distribution without side effects
Measurement of brain HBED levels after systemic administration confirmed its penetration in hippocampal mitochondria. - Executive summary:
Liang and colleagues investigated the distribution of HBED in rats (Liang, 2008). Chelatable iron is an important catalyst for the initiation and propagation of free radical reactions and implicated in the pathogenesis of diverse neuronal disorders. Studies have shown that mitochondria are the principal source of reactive oxygen species production after status epilepticus (SE). It was investigated whether SE modulates mitochondrial iron levels and whether consequent mitochondrial dysfunction and neuronal injury could be ameliorated with a cell-permeable iron chelator. Kainate (KA) induced SE resulted in a time-dependent increase in chelatable iron in mitochondrial but not cytosolic fractions of the rat hippocampus. Systemically administered N,N-bis (2-hydroxybenzyl) ethylenediamine-N,N-diacetic acid (HBED), a synthetic, BBB permeable iron chelator, ameliorated SE-induced changes in chelatable iron, mitochondrial oxidative stress (8-hydroxy-2 deoxyguanosine and glutathione depletion), mitochondrial DNA integrity and hippocampal cell loss. HBED significantly attenuates SE induced hippocampal neuronal damage. Measurement of brain HBED levels (brain bioavailability of HBED after systemic administration, measured in forebrain tissue and hippocampal mitochondria) after systemic administration of 75 µmol/kg, s.c. confirmed its penetration in hippocampal mitochondria. HBED levels were detected 1 h after administration, peaked at 3 h and remained stable for at least 6 h. Moreover, both tissues and mitochondrial HBED levels remained 25 % of the peak values until 24 h after injection. These results suggest that HBED penetrates the BBB and is accessible to hippocampal mitochondria. Total iron levels in the hippocampus were decreased 30% after 4 or 5 daily injections with 75 µmol/kg HBED. Increasing the injection frequency to 7 resulted in a 38% reduction of iron content compared with control animals but retarded weight gain. These results suggest a role for mitochondrial iron in the pathogenesis of SE-induced brain damage and subcellular iron chelation as a novel therapeutic approach (neuroprotective effects of HBED by inhibition of SE induced mitochondrial oxidative stress) for its management. However, HBED’s inability to influence behavioural seizure parameters produced by KA suggests that its mechanism of neuroprotection is not related to interference with acute seizure initiation and a direct antioxidant action of HBED cannot be ruled out.
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