[No public or meaningful name is available]

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vitro / ex vivo

Remarks:

transformation/dissolution in artificial physiological media

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2017-08-25 - 2018-01-26

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Objective of study:

bioaccessibility (or bioavailability)

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Series on Testing and Assessment No. 29 (23-Jul-2001): Guidance document on transformation/dissolution of metals and metal compounds in aqueous media

Deviations:

yes

Remarks:

Bioaccessibility testing: loading of 100 mg/L; five artificial physiological media agitated at 100 rpm, at 37 °C ± 2 °C; sampling after 2 h and 24 h; determination of Cr,Fe, Ti and Zn concentrations after filtration by ICP-OES.

Principles of method if other than guideline:

The test was performed on the basis of OECD Series on Testing and Assessment No. 29 as well as according to the bioaccessibility test protocol that has been developed on the basis of relevant published methods ([1], [2], [3], [4] and [5]).

The aim of this test was to assess the dissolution of the pigment IIPC-2018-010 (Titanium, iron and aluminium pseudobrookite and rutile) in the artificial physiological media GST, GMB, ALF, ASW and PBS. The test media were selected to simulate relevant human-chemical interactions (as far as practical), i.e. a substance entering the human body by ingestion or by inhalation.

Five different artificial physiological media with a single loading of test substance of 100 mg/Lwere used. The measurement of dissolved titanium, iron and aluminium concentrations after filtration were performed by ICP-OES. Samples were taken after 2 and 24 hours agitation (100 rpm) at 37 ± 2 °C. The study was performed in triplicate with two additional method blanks per medium.

[1] Hanawa T. 2004. Metal ion release from metal implants. Materials Science and Engineering C 24: 745-752.
[2] Stopford W., Turner J., Cappelini D., Brock T. 2004. Bioaccessibility testing of cobalt compounds. Journal of Environmental Monitoring 5: 675-680.
[3] Midander K., et al. 2007. In vitro studies of copper release from powder particles in synthetic biological media. Environmental Pollution 145: 51-59.
[4] European standard 1998. Test method for release of nickel from products intended to come into direct and prolonged contact with the skin (EN 1811)
[5] ASTM 2003. Standard test method for determining extractability of metals from art materials. ASTM D5517-03.

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

titanium, iron, aluminium oxide

Molecular formula:

x(Al, Fe)2TiO5 * yTiO2

IUPAC Name:

titanium, iron, aluminium oxide

Test material form:

solid: particulate/powder

Details on test material:

- Chemical description: Titanium, iron and aluminium pseudobrookite and rutile
- Physical state: Solid, brown powder, odourless
- Crystal structure: pseudobrookite and rutile
- Purity : ~ 97.9%
- Storage condition of test material: Kept dry in closed containers

Test animals

Species:

other: in vitro (simulated human body fluids)

Details on test animals or test system and environmental conditions:

Test principle in brief:
- five different artificial physiological media,
- single loading of test substance of 100 mg/L,
- samples taken after 2 and 24 hours agitation (100 rpm) at 37 ± 2 °C,
- two method blanks per artificial media were tested; measurement (by ICP-OES) of dissolved aluminium, iron and titanium after filtration
- the study was performed in triplicate

The aim of this test was to assess the dissolution of IPC-2018-010 (Titanium, iron and aluminium pseudobrookite and rutile) in five artificial physiological media: Artificial lysosomal fluid (ALF, pH = 4.5), Artificial sweat solution (ASW, pH = 6.5), Gamble´s solution (GMB, pH = 7.4), Artificial gastric fluid (GST, pH = 1.5), Phosphate buffered saline (PBS, pH = 7.4). The test media were selected to simulate relevant human-chemical interactions (as far as practical), i.e. a substance entering the human body by ingestion into the gastrointestinal tract and by inhalation.

Administration / exposure

Duration and frequency of treatment / exposure:

Samples were taken after 2 h and 24 h.

Doses / concentrations

Dose / conc.:

100 other: mg of the test item /L artificial media

Details on study design:

Reagents
The water (resistivity >18 MΩ·cm.) used for this test was purified with a Pure Lab Ultra water purification system from ELGA LabWater, Celle, Germany.
- Nitric acid - “Supra” quality (ROTIPURAN® supplied by Roth, Karlsruhe, Germany).
- Hydrochloric acid – “instra-analyzed plus” quality (J.T. Baker, Griesheim, Germany).
- Sodiumhydroxide – pro Analysis quality (Chemsolute, Th. Geyer, Renningen, Germany)

Metal analysis
- Standards:
- Certified reference materials: As quality control standards certified aqueous reference material TM-26.4 (lot no. 1115) and TMDA-52.4 (lot no. 0915) obtained from Environment Canada and a multielement standard (Roth Multielemenstandard, lot no. F38770, Karlsruhe, Germany) were analysed for total dissolved chromium, iron, titanium and zinc by ICP-OES along with the samples to determine the accuracy of the applied analytical method. Furthermore, the calibration solutions were measured along with the ICP-OES measurements as recalibration standards.

Instrumental and analytical set-up for the ICP-OES instrument:
Agilent 720, Agilent Technologies, Waldbronn, Germany
Nebulizer: Sea spray nebulizer from Agilent
Spray chamber: Glass cyclonic spray chamber from Agilent
Plasma stabilization time: at least 30 min before start of the measurements
Plasma gas flow: 15.0 L/min
Additional gas flow: 1.50 L/min
Carrier gas flow: 0.75 L/min
RF power: 1200W
Stabilization time of sample: 15 sec
Repetition time (three internal measurements per sample): 30 sec

Wavelengths: Al: 167.019 nm, 394.401 nm and 396.152 nm
Fe: 238.204 nm, 241.052 nm and 259.837 nm
Ti: 337.280 nm, 368.520 nm and 376.132 nm


The applied LOD/LOQ calculations for the Agilent 720 ICP-OES are (according to DIN 32645):
LOD: 3 x standard deviation of calibration blank/slope of the calibration
LOQ: 3 x LOD

Calibration: blank, 1 µg/L, 2 µg/l, 4 µg/L, 6 µg/L, 8 µg/L, 10 µg/L, 20 µg/L, 40 µg/L, 60 µg/L, 80 µg/L, 100 µg/L, 120 µg/L, 140 µg/L, 150 µg/L, 160 µg/L, 180 µg/L, 200 µg/L, 250 µg/L, 300 µg/L, 350 µg/L, 400 µg/L, 450 µg/L and 500 µg/L, for measurements of test vessel samples and for mass balance sample calibration was extended to 750 µg/L and 1000 µg/L.
Correlation coefficients (r): at least 0.995955

Details on dosing and sampling:

Loading:
The nominal loading in this test was 100 mg/L. However, due to weighing uncertainties the actual loadings range from 100.000 mg/L to 100.178 mg/L in the 15 test vessels.

Results and discussion

Main ADME results

Type:

other: Bioaccessibility

Results:

Higest dissolution at a loading of 0.1g/L in GST after 24h: Al = 179µg/L, Fe = 215µg/L, Ti =162µg/L

Bioaccessibility (or Bioavailability)

Bioaccessibility (or Bioavailability) testing results:

Concentration of dissolved aluminium in artificial physiological media.
Total Al ± SD in sample vessels with method blank subtraction
GST 2h 87.4 ± 3.41 µg/L
GST 24h 179 ± 8.78 µg/L
GMB 2h 8.67 µg/L
GMB 24h 25.4 µg/L
ALF 2h 52.9 ± 0.41 µg/L
ALF 24h 108 ± 0.70 µg/L
ASW 2h < LOD
ASW 24h 27.5 ± 2.42 µg/L
PBS 2h 2.57 µg/L
PBS 24h 24.2 µg/L

Concentration of dissolved iron in artificial physiological media.
Total Fe ± SD in sample vessels with method blank subtraction
GST 2h 61.2 ± 10.3 µg/L
GST 24h 215 ± 38.2 µg/L
GMB 2h GMB 24h ALF 2h 26.5 ± 0.78 µg/L
ALF 24h 97.7 ± 1.04 µg/L
ASW 2h ASW 24h 11.6 ± 1.35 µg/L
PBS 2h 2.26 ± 1.21 µg/L
PBS 24h 8.03 ± 6.53 µg/L

Concentration of dissolved titanium in artificial physiological media.
Total Ti ± SD in sample vessels without method blank subtraction
GST 2h 36.2 ± 4.57 µg/L
GST 24h 162 ± 8.75 µg/L
GMB 2h < LOD/LOQ
GMB 24h < LOQ
ALF 2h 17.9 ± 0.66 µg/L
ALF 24h 75.5 ± 1.33 µg/L
ASW 2h 1.44 ± 0.33 µg/L
ASW 24h 20.2 ± 1.13 µg/L
PBS 2h < LOD
PBS 24h 2.62 ± 0.17 µg/L

Any other information on results incl. tables

Method validation summary (ICP-OES)

Limits of detection (LODs), limits of quantification (LOQs) and correlation coefficients (r) are given for each medium and metal measured:

 

ASW medium

date	Chosen wavelength [nm]	LOD (µg/L)	LOQ (µg/L)	correlation coefficient	Sum of quality assurance samples	Recovery of quality assurance samples [%]
September 20, 2017; measurement of ASW test samples (2h and 24h) and method blanks	Al: 167.019 Fe: 259.837 Ti: 368.520	Al: 0.356 Fe: 1.13 Ti: 0.068	Al: 1.07 Fe: 3.38 Ti: 0.204	Al: 0.999958 Fe: 0.999979 Ti: 0.999957	Al: 37 Fe: 37 Ti: 37	Al: 95 Fe: 100 Ti: 100

 

PBS medium

date	Chosen wavelength [nm]	LOD (µg/L)	LOQ (µg/L)	correlation coefficient	Sum of quality assurance samples	Recovery of quality assurance samples [%]
October 05, 2017; measurement of PBS test samples (2h and 24h) and method blanks	Al: 167.019 Fe: 238.204 Ti: 376.132	Al: 0.252 Fe: 0.350 Ti: 0.201	Al: 0.756 Fe: 1.05 Ti: 0.604	Al: 0.999675 Fe: 0.999932 Ti: 0.999646	Al: 40 Fe: 40 Ti: 40	Al: 100 Fe: 100 Ti: 85

 

GMB medium

date	Chosen wavelength [nm]	LOD (µg/L)	LOQ (µg/L)	correlation coefficient	Sum of quality assurance samples	Recovery of quality assurance samples [%]
December 15, 2017; measurement of GMB test samples (2h and 24h) and method blanks	Al: 167.019 Fe: 238.204 Ti: 368.520	Al: 0.237 Fe: 0.432 Ti: 0.139	Al: 0.710 Fe: 1.29 Ti: 0.416	Al: 0.999848 Fe: 0.999860 Ti: 0.999778	Al: 28 Fe: 28 Ti: 28	Al: 100 Fe: 100 Ti: 100

 

GST medium

date	Chosen wavelength [nm]	LOD (µg/L)	LOQ (µg/L)	correlation coefficient	Sum of quality assurance samples	Recovery of quality assurance samples [%]
October 11, 2017; measurement of GST test samples (2h and 24h) and method blanks	Al: 396.152 Fe: 238.204 Ti: 376.132	Al: 0.953 Fe: 0.411 Ti: 0.944	Al: 2.86 Fe: 1.23 Ti: 2.83	Al: 0.999938 Fe: 0.999943 Ti: 0.999954	Al: 40 Fe: 40 Ti: 40	Al: 97.5 Fe: 100 Ti: 97.5

 

ALF medium

date	Chosen wavelength [nm]	LOD (µg/L)	LOQ (µg/L)	correlation coefficient	Sum of quality assurance samples	Recovery of quality assurance samples [%]
October 11, 2017; measurement of ALF test samples (2h and 24h) and method blanks	Al: 167.019 Fe: 259.837 Ti: 376.132	Al: 0.466 Fe: 1.22 Ti: 0.463	Al: 1.40 Fe: 3.66 Ti: 1.39	Al: 0.999978 Fe: 0.999958 Ti: 0.999953	Al: 40 Fe: 40 Ti: 40	Al: 100 Fe: 85.0 Ti: 100

  

Solution pH values

The target pH in all media before addition of test substance was in the nominal range.

During the study, the pH of GST, ALF and PBS media remained stable in the method blank vessels and the test vessels. Therefore, a possible effect of the test substance can be excluded.

In GMB medium, the pH in all vessels (including method blanks) increased during the time of the test from 7.44 / 7.45 /7.46 to 8.78, 8.84 and 8.96 (test vessels) and 7.46 to 8.79 and 8.84 (method blank vessels). Therefore, an effect of the test substance can be excluded. In fact, the pH of the GMB media does not seem to be stable under the conditions of the test.

In ASW medium, the pH in all vessels (including method blanks) decreased during the time of the test from 6.50 to 5.92 and 5.94 (test vessels) and 6.50 to 5.89 and 5.96 (method blank vessels). Therefore, an effect of the test substance can be excluded.

Temperature control

The test was performed in an incubated laboratory shaker (Shaking incubation cabinet, Minitron, INFORS AG, Bottmingen, Switzerland) at 100 rpm. The temperature was adjusted to 37.5 °C in a thermostatically controlled shaking cabinet to reach a temperature of 37 °C ± 2 °C in the media. The temperature remained stable during the test in all media.

Fortification

Mean recovery of fortified samples: 52.7 - 110 % (Al fortification), 87.7 - 107 % (Fe fortification), and 89.2 – 104% (Ti fortification)

Applicant's summary and conclusion

Conclusions:

On the basis of OECD Series on Testing and Assessment No. 29 as well as according to a bioaccessibility test protocol, which has been developed on the basis of relevant published methods, the dissolution of the pigment IPC-2018-010 (Titanium, iron and aluminium pseudobrookite and rutile) in the artificial physiological media (GST, GMB, ALF, ASW and PBS) with a single loading of 100 mg/L, agitation (100 rpm) at 37 °C ± 2 °C and sampling after 2 and 24 h, was determined. The measurement of dissolved aluminium, iron and titanium concentrations after filtration were performed by ICP-OES. The study was performed in triplicate with two additional method blanks per medium.

*Concentration of dissolved aluminium in artificial physiological media.
Total Al ± SD in sample vessels with method blank subtraction
GST 2h 87.4 ± 3.41 µg/L
GST 24h 179 ± 8.78 µg/L
GMB 2h 8.67 µg/L
GMB 24h 25.4 µg/L
ALF 2h 52.9 ± 0.41 µg/L
ALF 24h 108 ± 0.70 µg/L
ASW 2h < LOD
ASW 24h 27.5 ± 2.42 µg/L
PBS 2h 2.57 µg/L
PBS 24h 24.2 µg/L

Concentration of dissolved iron in artificial physiological media.
Total Fe ± SD in sample vessels with method blank subtraction
GST 2h 61.2 ± 10.3 µg/L
GST 24h 215 ± 38.2 µg/L
GMB 2h GMB 24h ALF 2h 26.5 ± 0.78 µg/L
ALF 24h 97.7 ± 1.04 µg/L
ASW 2h ASW 24h 11.6 ± 1.35 µg/L
PBS 2h 2.26 ± 1.21 µg/L
PBS 24h 8.03 ± 6.53 µg/L

Concentration of dissolved titanium in artificial physiological media.
Total Ti ± SD in sample vessels without method blank subtraction
GST 2h 36.2 ± 4.57 µg/L
GST 24h 162 ± 8.75 µg/L
GMB 2h < LOD/LOQ
GMB 24h < LOQ
ALF 2h 17.9 ± 0.66 µg/L
ALF 24h 75.5 ± 1.33 µg/L
ASW 2h 1.44 ± 0.33 µg/L
ASW 24h 20.2 ± 1.13 µg/L
PBS 2h < LOD
PBS 24h 2.62 ± 0.17 µg/L