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Biodegradation

The biodegradability of charcoal (Probe 2: C-Fix = 80.5%) was tested according to OECD guideline No. 301B (CO2 evolution test) and under GLP compliance.

The test item was tested in a concentration of 25 mg/L. As positive control, aniline was tested in parallel. The biodegradation of the test substance was determined by measuring the emitted CO2concentrations on test days 2, 4, 7, 9, 11, 14, 18, 23, during the 28-day test period and on day 29. The biodegradation for the whole test period was determined as <2%. Hence, charcoal (Probe 2: C-Fix = 80.5%) is considered not readily biodegradable. For the positive control, a degradation of 64% was achieved within 9 d, ensuring the validity of the test.

Adsorption/desoprtion screening

The adsorption coefficient KOCof charcoal (Probe 2: C-Fix = 80.5%) was determined using the HPLC method in accordance with the OECD guideline 121 and in compliance with GLP standards.

The study was performed using HPLC with a cyanopropyl chemical bound resin on a silica based column as stationary phase (pH = 5.5; 25 °C). As mobile phase methanol/water (55/45%, v/v) was used. Seven reference items with different retention times (tR) were chosen for the calibration on behalf of the retention time of the test item charcoal (Probe 2: C-Fix =80.5%) on the column.

First, a solution of the reference items was measured three times with HPLC, then a solution of the test item (three measurements), and finally the solution of the reference items was again measured three times.

For each reference item, the capacity factor k’ was calculated from the retention time of an unretained compound (sodium nitrate) and the retention time of the respective reference item. A calibration function was set up using the literature values for KOCof the reference items and the mean capacity factor of the six determinations. Based on the data set log k’ versus log KOC,the equation for the regression line was calculated.

The chromatogram of the test item showed one peak (mean TR= 2.254 min).The log KOCwas calculated as 0.661±0.026 (mean ± standard deviation) with a range of 0.633–0.686. As this log KOC-value lies outside the calibration range obtained with the used reference substances (1.32-5.63) and also outside the determinable log KOC-range of the HPLC screening method (according to OECD guideline 121: 1.5-5.0), the log KOCof the test item charcoal (Probe 2: C-Fix = 80.5%) should be stated as < 1.5.

Due to the limited solubility of the test item charcoal (Probe 2: C-Fix = 80.5%) in the solvent an extract instead of a solution was used for the measurements. Therefore, the results are necessarily related to the used charcoal extract with a nominal concentration of 208,910 mg/L in methanol/water (75/25% v/v) under the testing conditions used (25±1°C; pH 5.5).

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