Calcium bis[4-[[1-[[(2-chlorophenyl)amino]carbonyl]-2-oxopropyl]azo]-3-nitrobenzenesulphonate]

Specific investigations: other studies

Administrative data

Endpoint:

specific investigations: other studies

Remarks:

Biodurability and biodissolution in phagolysosomal simulant fluid

Type of information:

experimental study

Remarks:

study currently ongoing

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to

Guideline:

other: ISO:TR19057:2017

Version / remarks:

(Nti 2017), Nanotechnologies - Use and application of acellular in vitro tests and methodologies to assess nanomaterial biodurability

Test guideline 2

Qualifier:

equivalent or similar to

Guideline:

other: OECD draft TG on “solubility in aqueous media”

Version / remarks:

restricted to the „screening method“

Principles of method if other than guideline:

- Principle of test: The solubility of the organic pigment is determined by a static and a dynamic dissolution test after pre-treatment steps for purification of the samples.
For the static solubility experiment, the test material was suspended in a pH 4.5 phagolysosomal simulant fluid in a Schott glass bottle and shaked automatically for 24 hours after pre-treatment. The sample was analyzed by IR after combustion of its total organic content and specific fuorescence spectra or automated counting by SEM.
For the dynamic dissolution kinetic experiment, a continuous flow system was applied with the test material captured in a flow cell. The tempered medium was regulated at a constant flowrate by a pump at 37°C for 7 days and the fluid collection was performed with an autosampler. The particle size was analyzed by TEM as the median particle size before and after continuous flow (by manual evaluation of approximately 300 particles).

GLP compliance:

no

Type of method:

other: in chemico

Endpoint addressed:

basic toxicokinetics

repeated dose toxicity: inhalation

Test material

Reference

Name:

Unnamed

Type:

Constituent

Test material form:

solid

Administration / exposure

Route of administration:

other: test material is in direct contact to the phagolysosomal simulant fluid

Vehicle:

other: phagolysosomal simulant fluid

Remarks:

sodium phosphate dibasic anhydrous (Na2HPO4) 142.0mg/l; sodium chloride (NaCl) 6650 mg/l; sodium sulfate anhydrous (Na2SO4) 71 mg/l; calcium chloride dihydrate (CaCl2. 2H2O) 29 mg/l

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

UV-VIS spectrophotometry was used to analyze the extraction solvents in methanol/toluol (80:20), octanol and methanol. The general procedure is following “UV-VIS Absorption Spectra (spectrophotometric method)”, OECD guideline for Testing of Chemicals, guideline 101, adopted 12th May 1981.
Setting
Spectrophotometer: Perkin-Elmer Lambda 35
Wavelength range: 350-700 nm
Cell type: quartz, 10.0 mm
Data interval: 1.0 nm
Measuring mode: Absorbance
Slit: 1.0 nm
Scan speed: 120 nm/min
Smooth: 0 nm
Temperature: 25 °C

Calculation of the molar absorption coefficient
The molar absorption coefficient ε was calculated for absorption maxima >190 nm.
The formula for this calculation is ε = A/d * ci
ε = molar absorption coefficient [L mol-1cm-1]
A = Absorption
d = path length [cm]
ci = molar concentration of the test solution
= c / MM * 1000 [mol/L]
where
MM = molecular weight of test substance in g /mol =>MM = 459.84
c = concentration of the test substance in mg/L

Reference
Solvent (baseline/reference beam): the corresponding blank solutions were used

Duration of treatment / exposure:

- static solubility experiment: 24 hours
- dynamic dissolution kinetic experiment: 7 days

Frequency of treatment:

- static solubility experiment: one time application of medium
- dynamic dissolution kinetic experiment: continuous flow of medium

Post exposure period:

To avoid false positive results (detection of additives, impurities ect.) pigments are purified by sequential solvent washes: first Methanol/Toluol (80/20, Merck, HPLC grade), then N-Octanol (Sigma-Aldrich, Spectrophotometric grade), finally Methanol (Merck, HPLC grade).
For each solvent the pigment is shaken for 2 hours at room temperature, recovered by centrifugal pelletting (20,000rpm, 1h), and dried under vacuum (100 torr, 90°C, 1h). The solvent extracts are analyzed by UVVis spectroscopy and discarded.

Details on study design:

static solubility experiment:
- 10 mg of test substance suspended in 80 mL of medium (resulting concentration of 125 mg/L)
- text concentration is an order of magnitude higher than advised for metal oxide nanomaterials in the OECD draft TG on solubility in aquatic media (2017), because the LoD for organics by TOC is higher than for metals by ICPMS (which is the target of the OECD draft TG)
- closed bottle was placed in an IKA shaker to prevent settling
- after incubation, medium is filtered through a 1µm glasfilter directly followed by a 0.02µm (=20nm) aluminum based membrane (inorganic filters)
- after significant organic leaching by the 3kDa filters (as recommended by the OECD draft TG), this set-up was rejected

dynamic dissolution kinetic experiment:
- implementaiton of the Continous Flow Systm (ISO19057:2017), described by Koltermann-Juelly et al 2018
- 1 mg solids per flow cell applied in a single flow-cell
- up to 14 eluate samplings were taken