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Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013-02-26 to 2013-04-15
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report date:
2013

Materials and methods

Objective of study:
bioaccessibility (or bioavailability)
Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
The aim of this test was to assess the dissolution of metallic boron in the set of artificial physiological media. The test media were selected to simulate relevant human-chemical interactions (as far as practicable), i.e. contact of test substance with skin, a substance entering the human body by inhalation or by ingestion.
For the experimental setup the test item was weighed into flasks, adjusted to volume with the respective artificial physiological medium (loading of 100 mg/L) and agitated at 100 rpm at 37°C ± 2 °C. Samples were taken after 2 h and 24 h. The total dissolved boron concentrations of sampled solutions were determined after filtration (0.2 µm, Supor membrane) by ICP-OES.
GLP compliance:
yes (incl. QA statement)
Remarks:
signed, 2011-02-07

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
Test item name: Metallic boron
Molecular formula: B
Molecular weight: 10.812 g/mol
State of matter and appearance: grey powder, odourless
Storage conditions: Store dry in tightly closed containers. Avoid high humidity
Stability: Stable under storage conditions



Radiolabelling:
no

Test animals

Species:
other: in vitro (simulated human body fluids)
Details on test animals or test system and environmental conditions:
Five different artificial physiological media, single loading of test substance of 100 mg/L, measurement of dissolved boron concentrations after 2 and 24 hours agitation (100 rpm) at 37 ± 2°C, two additional method blanks per medium. The study was performed in triplicate.
The total dissolved boron concentrations of sampled solutions were determined after filtration (0.2 µm, Supor membrane) by ICP-OES.

Administration / exposure

Details on dosing and sampling:
For the experimental part, three independent flasks (polycarbonate) per media were prepared with a loading of 100 mg/L.
Two additional control blanks (same procedure) were also prepared. Three replicates and two method blanks per artificial media were tested; solutions were sampled after 2 and 24 h, to measure total dissolved boron concentrations (by ICP-OES), temperature and pH.
All samples were filtered through 0.2 µm filter (Syringe Filter w / 0.2 µm, polyethersulfon membrane, DIA Nielsen, Dueren, Germany) prior to further treatment.
Aqueous samples of approx. 20 mL taken for B analysis were transferred into disposable scintillation vials (20 mL scintillation tubes, Sarstedt, Nuembrecht, Germany), acidified (target conc. 3 % HNO3) and stored at approx. 4 °C until analysis.

Analysis of total dissolved boron in test and in mass balance samples by ICP-OES
The analysis of total dissolved boron concentrations in test and mass balance samples were measured using an Agilent 720 ICP-OES (Agilent Techologies, Waldbronn, Germany. Boron was detected at the wavelength 182.577 nm, 249.678 nm, 249.772 nm, 208.956 nm and 208.889 nm. The following solutions were used to calibrate the instrument: blank, 1 µg/L, 2.5 µg/L, 5 µg/L, 7,5 µg/L, 10 µg/L, 25 µg/L, 50 µg/L, 75 µg/L, 100 µg/L, 250 µg/L, 500 µg/L, 750 µg/L and 1000 µg/L. Calibrations were performed before each measurement. The calibration formula was calculated using the linear regression algorithm of the ICP-OES instrument. The respective wavelength data with the best recoveries for the validation samples (certified reference material and recalibration standards) in the measurement series and a correlation coefficient with at least 0.995 were used for calculating concentrations (i.e. 249.772 nm for boron). Correlation coefficients (r) for the wavelengths used for evaluation of data were at least 0.9999. For each sample, at least three internal measurements were performed and the mean was calculated and printed by the instrument software.

Six measurements were performed for the determination of boron concentration in the test item samples, method blanks, mass balance samples and filter samples.
The applied LOD/LOQ calculations for the Agilent 720 ICP-OES are (according to DIN 32645):
LOD: 3 * standard deviation of calibration blank/slope of the calibration
LOQ: 3 * LODThe certified reference materials TM-15.2 (certified with 23.1 µg/L boron) and TM-DWS.2 (certified with 81.5 µg/L boron) as well as the recalibration standard (100 µg B/L) were analyzed as quality assurance samples along with the test samples. To meet quality assurance requirements recovery needs to be in the range of ± 15 % of the respective certified value

For the QA/QC of ICP-OES measurements of boron test samples, the mean accuracy and precision were 106 ± 3.6 % (n = 18) for TM-15.2 and 102 ± 3.2 % (n = 7) for TM-DWS.2 (dilution factor 2) and 107 ± 1.2 % (n = 10) for TM-DWS.2 (no dilution factor). The average recovery for the recalibration standard 100 µg/L was 102 ± 3.7 % (n = 17).

From selected artificial physiological media, two samples were fortified with a known amount of boron (by standard addition of commercial standards) to determine the standard recovery of boron.For fortified test samples, recoveries were in the range 90.0 – 97.3 % for B.

Fortification of test samples.
sample measured calculated level recovery
concentration after addition
[µg/L] [µg/L] [µg/L] / [%]
Vessel A sample 1 GMB 2h 113 168 168 / 100
Vessel A sample 1 GMB 24h 131 179 171 / 95.5
Vessel A sample 1 ALF 2h 111 166 164 / 98.4
Vessel B sample 1 ALF 2h 110 166 163 / 98.0
Vessel B sample 1 GST 2h 172 203 202 / 99.2
Vessel C sample 1 GST 2h 176 205 205 / 100
Vessel A sample 1 PBS 2h 116 170 176 / 104
Vessel A sample 1 PBS 24h 139 183 185 / 101
Vessel A sample 1 ASW 2h 107 164 172 / 104
Vessel A sample 1 ASW 24h 127 176 182 /104

For the measurement series of the mass balance, the mean accuracy and precision were 107 ± 0.8 % (n = 6) for TM-15.2 (dilution factor 2), 105 ± 1.6 % (n = 6) for TM-15.2 (no dilution), 102 ± 1.3 % (n = 6) for TM-DWS.2 (dilution factor 10) and 103 ± 1.7 % (n = 6) for TM-DWS.2 (dilution factor 2). The average recovery for the recalibration standard 100 µg/L was 98.6 ± 0.9 % (n = 12).

From mass balance vessels as well as from filter samples, selected samples were fortified with a known amount of boron (by standard addition of commercial standards) to determine the standard recovery of boron. For fortified test samples, recoveries of B were in the range 98.8 – 99.5 % for mass balance vessel and 96.8 – 99.3 % for mass balance filters.

Results and discussion

Any other information on results incl. tables

Solution pH values

After preparation of the artificial physiological media, solution pH was adjusted to their respective target pH. Solution pH data, measured prior to the test, after 2h and 24h in the different media, are compiled infollowing table.

Measured pH in method blank and test vessels in artificial media:

Mean of

target pH

pH prior to the test

temp. [°C] prior to the test

pH after 2h

temp.[°C]

pH after 24h

temp.[°C]

ALF vessel A-C

4.50

4.50

30.20

4.50

33.37

4.43

29.43

ALF blank vessel A+B

4.50

4.50

30.65

4.40

36.50

4.40

34.00

ASW vessel A-C

6.50

6.50

32.90

5.53

34.37

5.50

31.83

ASW blank vessel A+B

6.50

6.50

33.85

5.45

36.60

5.35

35.40

GMB vessel A-C

7.40

7.40

31.60

8.80

33.73

9.10

31.23

GMB blank vessel A+B

7.40

7.40

31.80

8.80

36.45

9.00

34.75

GST vessel A-C

1.50

1.60

35.70

1.50

35.67

1.47

33.70

GST blank vessel A+B

1.50

1.60

35.50

1.50

36.75

1.50

36.30

PBS vessel A-C

7.30

7.30

34.97

7.20

34.83

7.20

32.93

PBS blank vessel A+B

7.30

7.30

35.00

7.20

36.40

7.20

36.30

The target pH in all media before addition of test substance is in the nominal range. During the study, the pH of ALF, GST and PBS solutions remained stable.

 

The pH of the ASW media in the sample vessels is first increasing to a pH of 7.3 (2 h sampling) and after this decreasing to a pH of 6.2 (24 h).In contrast the pH of ASW media in the method blanks is decreasing during the time of the test from 6.5 to 6.0 and 6.9. Therefore, an effect of the test substance cannot be excluded.

 

In GMB medium, the pH in all vessels (including method blanks) increased during the time of the test from 7.4 to 8.8 (test vessels) and 7.4 to 8.8 (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.

Analytical measurements The results of the ICP-OES measurements of total dissolved boron concentrations in samples, the resulting boron concentrations in method blanks and the mean dissolved total boron concentrations are presented in the attached document (Analytical measurments boron) in Table 1 - Table 5. The summary in given in following table:

Concentration of boron in artificial media.

media and sample

total B ± SD in method blanks [µg/L]

total B ±SD in sample vessels [µg/L]

B ± SD in sample vessels with blank subtraction [µg/L]

ALF 2h

3.99 ± 0.50

115 ± 5.32

111± 5.32

ALF 24h

4.20 ± 0.32

137 ± 7.38

133 ± 7.38

ASW 2h

12.6 ± 0.37

108 ± 3.96

96.4 ± 3.96

ASW 24h

11.3± 4.50

127 ± 5.23

115± 5.23

GMB 2h

21.5 ± 0.30

130 ± 26.4

108 ± 26.4

GMB 24h

22.5 ± 0.25

146 ± 25.2

124± 25.2

GST 2h

32.2 ± 2.23

167 ± 16.2

134 ± 16.2

GST 24h

34.2 ± 2.35

381 ± 58.4

348 ± 58.4

PBS 2h

3.45 ± 0.18

131 ± 27.0

127 ± 27.0

PBS 24h

3.89 ± 0.19

155 ± 26.2

151 ± 26.2

Mass balance calculation

Total dissolved boron concentrations in vessels, filters and syringes as measured by ICP-OES indicate a complete dissolution of metallic boron in all physiological media after addition of aqua regia to the sample vessels.

Concentration of boron in one mass balance sample (vessel and filter / syringes) for each media.

media

value for dissolved B

after addition of

aqua regia
[mg]

nominal
concentration
[mg] #

recovery
[%]

ALF vessel B

48.7

48.4

101

ASW vessel A

50.8

48.8

104

GMB vessel B

50.9

48.8

104

GST vessel A

50.3

48.4

104

PBS vessel B

50.2

48.4

104

# nominal concentration B = 95.9 % in test item -> 47.95 mg B in 50 mg test item -> 47.95 * initial weight / 50mg

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): other: Dissolved boron concentration under physiologically relevant conditions were between ca. 100 and 200 µg/L (0.1-0.2% of material dissolved) in all media.
The bioaccessibility of elemental boron powder has been investigated experimentally in vitro by simulating dissolution in artificial physiological media considered to mimic the most relevant exposure routes (oral, dermal and inhalation) at a loading of 100 mg/L. In conclusion, dissolved boron concentrations under physiologically relevant conditions were between ca. 100 and 200 µg/L (0.1-0.2% of material dissolved) in all media.
Executive summary:

The bioaccessibility of boron metal powder has been investigated experimentally in vitro by simulating dissolution in artificial physiological media considered to mimic the most relevant exposure routes (oral, dermal and inhalation).

Five media were used:

- Gamble’s solution (GMB, pH 7.4) which mimics the interstitial fluid within the deep lung under normal health conditions,

- phosphate-buffered saline (PBS, pH 7.2), which is a standard physiological solution that mimics the ionic strength of human blood serum,

- artificial sweat (ASW, pH 6.5) which simulates the hypoosmolar fluid, linked to hyponatraemia (loss of Na+ from blood), which is excreted from the body upon sweating,

- artificial lysosomal fluid (ALF, pH 4.5), which simulates intracellular conditions in lung cells occurring in conjunction with phagocytosis and represents relatively harsh conditions and

- artificial gastric fluid (GST, pH 1.5), which mimics the very harsh digestion milieu of high acidity in the stomach.

In conclusion, dissolved boron concentration under physiologically relevant conditions were between ca. 100 and 200 µg/L (0.1-0.2% of material dissolved) in all media.