[No public or meaningful name is available]

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reason / purpose for cross-reference:

read-across source

Duration:

3 h

Dose descriptor:

IC10

Effect conc.:

6 000.2 other: ppm

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: 5573.6 - 6426.8 ppm

Key result

Duration:

3 h

Dose descriptor:

NOEC

Effect conc.:

1 000 other: ppm

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No data available.

- Effect concentrations exceeding solubility of substance in test medium: no effects observed for all tested concentrations.

Results with reference substance (positive control):

- Results with reference substance valid? Yes.

- Relevant effect levels: 3-Hour EC50 value was determined to be 15.5 mg/L.

Reported statistics and error estimates:

Analysis of Consumption Data:

Only the linear section on the chart paper representing the oxygen consumption was used to evaluate the results. The chart readings were transformed into mg of O2 per liter consumed per hour (mg O2/L*hr). To calculate the two control respiration rates for each test material concentrations:
1 ─ 2 *100* RS/(Rc1+Rc2),

Where:
RS = oxygen consumption rate of the inoculum exposed to the test material;

Rc1 = oxygen consumption rate, control 1;

Rc2 = oxygen consumption rate, control 2.

In addition to the assessment of the test material, the reference toxicant results were similarly evaluated. The reference toxicant data were subject to probit analysis to calculate the EC50.

To calculate the NOAEC, a t test was performed.

Consumption data were evaluated following the guidelines of USEPA(1989) in order to estimate the concentration which would produce a10%inhibition of oxygen consumption. The estimate is produced using a linear interpolation and requires that the inhibition increases monotonically with increasing exposure concentration. If this condition is not met, the data are smoothed to satisfy the condition.

Because the test concentrations were not replicated, the Jackknife statistical method was used to calculate a variance and the 95% confidence interval for the IC10. This method consists of calculating an IC10 using the first control value, then the second control value and finally the mean of the two control values. Additionally, each test concentration is dropped from the calculation in turn. This results in a number of IC10 estimates. The variance in these resultant IC10 estimates is used to produce a confidence interval with the degree of freedom based on the number of estimates.

Table 2: Oxygen consumption and Chemical-Induced Inhibition of Bacterial Respiration

Chemical Conc.  (replicate #)	O2Consumption (mg/L*hr)	Percent Inhibition
Negative controls
Sample 1	54.7	─
Sample 2	55.4	─
Test material
1 ppm (1)	59.0	-7.2
10 ppm (1)	50.7	7.9
100 ppm (1)	61.5	-11.7
1000 ppm (1)	66.4	-20.6
10,000 ppm (1)	47.2	14.3
Positive control
10 ppm	41.1	24.8
20 ppm	16.4	70.2
40 ppm	6.2	88.7

Statistical analysis of the results

The resultant respiration rates were respectively: 59.0, 50.7, 61.5, 66.4, and 47.2 mg O₂/L*hr after treated with 1, 10, 100, 1000 and 10000 ppm test material. These data translate into percent inhibitions of -7.2, 7.9, -11.7, -20.6, and 14.3 %. Because none of the test concentrations produced an inhibition greater than 50%, an EC₅₀could not be calculated. Each respiration rate was statistically compared as a single value versus the mean of the controls following the of Sokal & Rohlf (1981) using a single-sidedttest. The results of thettest revealed that the 10,000 ppm test concentration expressed a statistically significant inhibition of oxygen consumption as compared to the controls. The discriminatory power of the test is low, however, due to having only one degree of freedom.

An IC₁₀was calculated using a point estimate technique and Jackknifed to produce a variance from which a confidence could be derived. The result estimate for the IC₁₀was 6000.2 ppm of the test material. The 95% confidence interval for this estimate is from 5573.6 to 6426.8 ppm.

Validity criteria fulfilled:

yes

Conclusions:

The calculated no-observed-adverse-effect concentration (NOAEC) based on inhibition compared to the control was calculated to equal 1000 ppm. The EC50 could not be calculated. The IC10 was calculated to equal 6000.2 ppm with a 95 % confidence interval of 5573.6 to 6426.8 ppm.

Executive summary:

The potential impact of test material on microbial metabolism, as represented by the consumption of oxygen, was investigated using the "Activated Sludge, Respiration Inhibition Test" as prescribed by OECD (1984) and detailed in WCC Protocol OECD209 (Expanded Range Procedures). The test duration was a three-hour exposure period to the test material followed by up to ten minutes for the measurement of oxygen consumption. The study design was comprised of five nominal exposure concentrations: 1, 10, 100, 1000 and 10,000 ppm; a duplicate control group; and an assessment of the sensitivity of the inoculum used in the test to a reference toxicant (3,5-dichlorophenol).

The activated sludge respiration test with test material passed the quality control criteria for an acceptable test. The EC₅₀calculated for the reference toxicant was 15.5 rng/L, within the acceptable range of 5 to 30 mg/L. The two control replicates produced oxygen consumption rates within the required 15 % of each other, 54.7 and 55.4 mg O₂/L*hr.

The respiration rates of the sludge-associated microbes exposed to the five nominal concentrations of test material were 59.0, 50.7, 61.5, 66.4 and 47.2 mg O₂/L*hr respectively. The calculated NOAEC based on inhibition compared to the control calculated to equal 1000 ppm. The EC₅₀ for this material regarding the inhibition of metabolism, as represented by respiration, was greater than 10,000 ppm, the IC₁₀was calculated to equal 6000.2 ppm with a 95 % confidence interval of 5573.6 to 6426.8 ppm.

Description of key information

The calculated no-observed-adverse-effect concentration (NOAEC) based on inhibition compared to the control was calculated to equal 1000 ppm. The EC50 could not be calculated. The IC10 was calculated to equal 6000.2 ppm with a 95 % confidence interval of 5573.6 to 6426.8 ppm.

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:

1 000 mg/L

Additional information

The potential impact of test material on microbial metabolism, as represented by the consumption of oxygen, was investigated using the "Activated Sludge, Respiration Inhibition Test" as prescribed by OECD (1984) and detailed in WCC Protocol OECD209 (Expanded Range Procedures). The test duration was a three-hour exposure period to the test material followed by up to ten minutes for the measurement of oxygen consumption. The study design was comprised of five nominal exposure concentrations: 1, 10, 100, 1000 and 10,000 ppm; a duplicate control group; and an assessment of the sensitivity of the inoculum used in the test to a reference toxicant (3,5-dichlorophenol).

The activated sludge respiration test with test material passed the quality control criteria for an acceptable test. The EC₅₀calculated for the reference toxicant was 15.5 rng/L, within the acceptable range of 5 to 30 mg/L. The two control replicates produced oxygen consumption rates within the required 15 % of each other, 54.7 and 55.4 mg O₂/L*hr.

The respiration rates of the sludge-associated microbes exposed to the five nominal concentrations of test material were 59.0, 50.7, 61.5, 66.4 and 47.2 mg O₂/L*hr respectively. The calculated NOAEC based on inhibition compared to the control calculated to equal 1000 ppm. The EC₅₀for this material regarding the inhibition of metabolism, as represented by respiration, was greater than 10,000 ppm, the IC₁₀was calculated to equal 6000.2 ppm with a 95 % confidence interval of 5573.6 to 6426.8 ppm.