Lithium nickel potassium oxide

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

STUDY COMPLETED
08 Feb 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Commission Regulation (EC) No 2016/266, C.3 OECD Guideline for Testing of Chemicals, No. 201

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Remarks:

The results from KDLNO are compared to 3,5 Dichlorphenol EC50 values published in United Kingdom Environment Agency (2006) Monitoring Certification Scheme (MCERTS) Performance Standard for Laboratories Undertaking Direct Toxicitiy Assessment of Effluent.

Vehicle:

no

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

Insertion of test organisms
The cell density of the inoculum culture in exponential growth phase was determined (Section
3.3.1) and then adjusted to 30 x 104 cells/mL. To obtain the correct initial cell density in the test
replicates of 0.3 x 104 cells/mL and the correct nominal concentrations of the test solutions,
inoculum culture was added to each test solution at a ratio of 1:100.
The initial inoculum biomass is reduced at start of exposure in this test due to a mistake in the
calculation for the inoculum

Test type:

static

Water media type:

other: The test medium (OECD medium) was prepared according to OECD Guideline for Testing of Chemicals, No. 201 Algal growth inhibition test. The pH value of the test medium was 7.8 and was adjusted to pH 8.1 by the addition of 1M NaOH.

Remarks:

The test medium (OECD medium) was prepared according to OECD Guideline for Testing of Chemicals, No. 201 Algal growth inhibition test. The pH value of the test medium was 7.8 and was adjusted to pH 8.1 by the addition of 1M NaOH.

Limit test:

no

Total exposure duration:

72 h

Test temperature:

23.7 – 23.8 °C

pH:

pH-values at the start (t = 0 h) and at the end (t = 72 h) of the exposure

Nominal and measured concentrations:

TEST GROUPS AND CONCENTRATIONS
Test groups: 0 (control), 0.32, 1.0, 3.2, 10, 32 and 100 mg/L as loading rate based on test substance mass without correction for purity.
Test replicates:
6 replicates for the Control;
3 replicates for each test substance concentration
1 additional replicate per test group for initial concentration control analysis only.
1 additional uninoculated (without algae) replicate per test group for background fluorescence correction and to determine the influence of the test design on test substance stability.

Reason for selection of test concentrations:
According to the test guidelines, at least 5 concentrations in a geometric series with a separation factor of ≤3.2 should be used, preferably encompassing the range from 5 – 75% inhibition of algal growth.

The test concentrations were selected on the basis of a range finding test (experimental conduct in accordance with GLP but without a GLP Status). The results of the 72 hour range finding
test were (as nominal concentrations):
EfluorescenceC50 = between 10 and 100 mg/L (18 mg/L estimated);
ErC50 = between 10 and 100 mg/L (39 mg/L estimated);
Pretest results showed a shallow concentration response algal growth curve ranging from 2% inhibition at 1 mg/L to 79% inhibition at 100 mg/L. Due to the flat response curve, this test
was performed using a broad concentration range down to 0.32 mg/L.

Details on test conditions:

Test conditions
Test duration: 72 hours
Test vessels: Erlenmeyer flasks (nominal volume 250 mL) plugged with gas
permeable silicone sponge caps
Test media: OECD media
Test volume: 100 mL
Initial cell density: 0.3 x 104 cells/mL
Test chamber: Vötsch Industrietechnik GmbH Bioline (VB1014) controlled
climate cabinet.
Test temperature: 23.7 – 23.8 °C
Illumination: Artificial light, type universal white (OSRAM L 25), permanent illumination. To minimize the potential effect of slight variations
in illumination, the test vessels were rearranged daily.
Light intensity: 6882 lux (within ± 15% variability) at a wave length of 400 - 700 nm
Shaking rate: Continuous (approx. 85 rpm)
Test parameter: Algal growth measured as in vivo chlorophyll-a fluorescence
(pulsed excitation with light flashes having a wavelength of 430 nm)
Route of exposure: Static exposure via the test medium.


Insertion of test organisms
The cell density of the inoculum culture in exponential growth phase was determined and then adjusted to 30 x 104 cells/mL. To obtain the correct initial cell density in the test
replicates of 0.3 x 104 cells/mL and the correct nominal concentrations of the test solutions,
inoculum culture was added to each test solution at a ratio of 1:100.
The initial inoculum biomass is reduced at start of exposure in this test due to a mistake in the
calculation for the inoculum.


OBSERVATIONS AND MEASUREMENTS
pH, fluorescence, and test substance:
Measurements and samples taken accordingly as guided by test protocols

Fluorescence measurement:
The fluorescence of aliquots from each test vessel was measured using a Tecan Infinite 200Pro fluorometer in a 96- well flat bottom black plate with the following parameters:
fluorescence top reading; excitation / emission wavelength = 430 / 670 nm; excitation / emission bandwidth = 20 / 25 nm; flashes = 5; integration time = 20 μs; shaking duration = 15s;
shaking amplitude = 6 mm. After measurement, aliquots were discarded.

Temperature measurement:
Continuous measurement during the whole test period in the climate chamber in a separate deionized water filled flask.

Determination of correlation between fluorescence and cell density:
After the end of the exposure the control replicates were mixed and serially diluted by factor 2. The fluorescence of aliquots from the undiluted mixture and the dilutions were measured and
in parallel cell density was determined by a direct microscopic count (two counts in a Neubauer haemocytometer). These data were used to derive a linear correlation between fluorescence
and cell density.

Algal morphology: The inoculum culture was observed microscopically at the start of the test to verify normal and healthy cells. At test termination, a pooled sample from each test concentration was examined and any abnormal appearance of the algae noted.

Light measurement: Light homogeneity was evaluated by measuring light intensity at 5 locations within the incubation area at the start and end of the test (Testo 545 light meter, Testo GmbH & Co, Lenzkirch, Germany). The light intensity did not vary by more than ± 15% over the incubation area.

Data Evaluation

All reported data values are rounded to the appropriate significant figures based on the precision of the analytical method and/or consistent with the requirements in the pertinent test guideline(s).

The mean fluorescence was determined after 0, 24, 48, 72 h and converted to cell density
using a correlation curve derived from a microscopic count of the control.
The commercial software "TOXRAT Professional 2.10” (ToxRat Solutions GmbH, Alsdorf,
Germany) was used for the statistical evaluation of the data. The yield and specific growth rate
over the exposure period was calculated for each replicate flask of each test group and
inhibition for each test group was determined by comparison to the control. The percent
inhibition of the mean yield and growth rate compared to the control was calculated for each
test group replicate. The data are illustrated using plots of percent inhibition (response) versus
concentration. ECx values and confidence limits were calculated by probit analysis. The LOEC was determined by comparing the means of the calculated yield or growth rate of the various concentration levels with the control using Dunnett’s multiple t-test (onesided).
The NOEC was the next tested concentration below the LOEC.

VALIDITY CRITERIA
This test was fully compliant with all the following validity criteria required by the corresponding
test guidelines and is considered valid.
• The validity criterion for cell multiplication factor in the untreated control is > 16 in 72 hours.
The cell multiplication factor in the untreated control (all replicates mixed together) was 396-
fold after 72 hours.
• The validity criterion for the mean coefficient of variation for section-by-section growth rates
for each test day in the control cultures is ≤35%. The mean coefficient of variation for
section-by-section growth rates for each test day in the control cultures was 18.1%.
• The validity criterion for the coefficient of variation of average specific growth rates during
the whole test period in replicate control cultures is ≤7%. The coefficient of variation of
average specific growth rates during the whole test period in replicate control cultures was
2.6%.

Reference substance (positive control):

yes

Remarks:

The results are compared to 3,5 Dichlorphenol EC50 values published in United Kingdom Environment Agency (2006) Monitoring Certification Scheme (MCERTS) Performance Standard for Laboratories Undertaking Direct Toxicitiy Assessment of Effluent

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

54.5 mg/L

Nominal / measured:

nominal

Conc. based on:

other: The aqueous fraction of the test solution, after separation of the undissolved material, was considered the water saturated fraction (WSF) in test media.

Basis for effect:

growth rate

Remarks:

Exponentially growing algae are cultured under defined conditions for several generations in the presence of the test substance, and cellular biomass over the exposure period is determined in relation to an untreated control.

Details on results:

In a 72-hour algal growth study, cultures of Pseudokirchneriella subcapitata were exposed to KDLNO at loading rates of 0 (control), 0.32, 1, 3.2, 10, 32 and 100 mg/L under static conditions in accordance with the OECD 201 guideline. The water pH, temperature and lighting intensity were all maintained within acceptable guideline specifications.
The following statistical effect concentrations were obtained after 72 hours of exposure:
ErC50/ErL50: 54.5 mg/L
confidence limits 95%: 31.9-93.1 mg/L

Since the test substance is a poorly water soluble inorganic chemical, only the dissolved content of the constituent element (nickel) was analytically determined. All reasonable efforts were taken to produce a saturated solution of the test substance in test media, following the guidance in OECD 23. According to OECD 23, for tests with chemicals that can not be quantified by analytical methods at the concentrations causing effects, the effect concentration can be expressed based on the nominal concentrations (the loading rate). According to the preliminary information of the Competence Center Analytics of BASF SE, the component (nickel) was measured at the tested loading rates ≥3.2 mg/L, which indicates that test item must have been present.

The results in this study are consistent with all validity criteria and the test is valid according to the guidelines of this study. No deviations from test guidelines or other incidents occurred during the course of the reported test which may have influenced the results.

Validity criteria fulfilled:

yes

Conclusions:

The following effect concentrations (mg/L) were obtained after 72-h based on loading rate:
ErC50/ErL50: 54.5 mg/L
confidence limits 95%: 31.9-93.1 mg/L

Executive summary:

In a 72-hour algal growth study, cultures of Pseudokirchneriella subcapitata were exposed to KDLNO at loading rates of 0 (control), 0.32, 1, 3.2, 10, 32 and 100 mg/L under static conditions in accordance with the OECD 201 guideline. The water pH, temperature and lighting intensity were all maintained within acceptable guideline specifications.

The following statistical effect concentrations were obtained after 72 hours of exposure:

ErC50/ErL50: 54.5 mg/L

confidence limits 95%: 31.9-93.1 mg/L

 

Since the test substance is a poorly water soluble inorganic chemical, only the dissolved content of the constituent element (nickel) was analytically determined. All reasonable efforts were taken to produce a saturated solution of the test substance in test media, following the guidance in OECD 23. According to OECD 23, for tests with chemicals that can not be quantified by analytical methods at the concentrations causing effects, the effect  concentration can be expressed based on the nominal concentrations (the loading rate). According to the preliminary information of the Competence Center Analytics of BASF SE, the component (nickel) was measured at the tested loading rates ≥3.2 mg/L, which indicates that test item must have been present. 

The results in this study are consistent with all validity criteria and the test is valid according to the guidelines of this study. No deviations from test guidelines or other incidents occurred during the course of the reported test which may have influenced the results.

 

Description of key information

In a 72-hour algal growth study, cultures of Pseudokirchneriella subcapitata were exposed to KDLNO at loading rates of 0 (control), 0.32, 1, 3.2, 10, 32 and 100 mg/L under static conditions in accordance with the OECD 201 guideline. The water pH, temperature and lighting intensity were all maintained within acceptable guideline specifications.

The following statistical effect concentrations were obtained after 72 hours of exposure:

ErC50/ErL50: 54.5 mg/L

confidence limits 95%: 31.9-93.1 mg/L

 

Since the test substance is a poorly water soluble inorganic chemical, only the dissolved content of the constituent element (nickel) was analytically determined. All reasonable efforts were taken to produce a saturated solution of the test substance in test media, following the guidance in OECD 23. According to OECD 23, for tests with chemicals that can not be quantified by analytical methods at the concentrations causing effects, the effect  concentration can be expressed based on the nominal concentrations (the loading rate). According to the preliminary information of the Competence Center Analytics of BASF SE, the component (nickel) was measured at the tested loading rates ≥3.2 mg/L, which indicates that test item must have been present. 

The results in this study are consistent with all validity criteria and the test is valid according to the guidelines of this study. No deviations from test guidelines or other incidents occurred during the course of the reported test which may have influenced the results.

 

Key value for chemical safety assessment

EC50 for freshwater algae:

54.5 mg/L