Lithium bis(oxalato)borate

Endpoint summary

Additional information

Short term toxicity to fish

LiBOB was assessed in a short-term toxicity to fish study, according to OECD guideline 203 and EU method C.1. Young zebrafish were exposed to the test item in a semi-static test for 96 h at nominal concentrations of 0, 100 and 150 mg/L. The test item showed no mortality up to a nominal concentration of 150 mg/L. Therefore the 96 h LC 50 was determined to be greater than 150 mg/L and the 96 h NOEC was determined to be 150 mg/L. (MEP, 2011)

Long term toxicity to fish

Under semi-static conditions of 48h-renewal, the effect on growth rates and other observed effects in juvenile fish (Gobiocypris rarus) exposed to the test substance LiBOB for 28 days was conducted according to the following guidelines: "The guidelines for the testing of chemicals" (HJ/T 153 -2004 ) and the OECD Guideline 215 "Fish, Juvenile Growth Test" (2000) and in compliance with GLP criteria.

Nominal concentrations of 0, 1.25%, 2.5%, 5%, 10% and 20% parent solutions were used in the test.

During the whole test period, the pH values of the control and test media were between 7.66 and 7.89, and the Dissolved Oxygen (DO) values varied from 92.7% ~ 96.9% of the air saturation at the test temperature, and the temperature of the test media was maintained at 22.9 °C ~ 23.2 °C.

All fishes in the control group were normal. The mean weight of fish in the control group increased more than 50%. So the study met the acceptability criteria prescribed by the protocol (dissolved oxygen concentration: no less than 60% of the air saturation value; temperature: (23±2) °C and not differ by more than ± 1°C between test chambers; the increasing rate of the mean wet-weight of fish: no less than 50% of the initial weight). Therefore the test is valid.

ICP-MS was used to determine to the concentration of the test substance in the test medium based on the Lithium ion. For the test solutions with the concentration of 1.25, 2.50, 5.00, 10.0 and 20.0 mg/L, the mean measured concentrations were 1.35, 2.50, 4.97, 9.92 and 19.6 mg/L, respectively. This indicated that the test concentration for the test solution (based on the lithium ion) was stable throughout a period of 48h (deviation within 20%). Thus a semi-static test of 48h-renewal was reasonable.

The test results showed that under valid semi-static test conditions ( 48h-renewal), all fish in the control groups and treated groups (1.25, 2.50, 5.00, 10.0 and 20.0 mg/L) were alive and appeared normal. In the 28d test period, the LOEC of the test substance for the juvenile fish

growth rate is greater than 20.0 mg/L (measured concentration was 19.6 mg/L), and NOEC is equal to20.0 mg/L (measured concentration was 19.6 mg/L), thus:

28 d-LOEC > 20.0 mg/L (measured concentration was 19.6 mg/L);

28 d-NOEC = 20.0 mg/L (measured concentration was 19.6 mg/L).(MEP, 2017)

Short-term toxicity to aquatic invertebrates

Lithium bis(oxalato)borate was assessed in a short term toxicity study to Daphnia magna according to OECD 202 and EU method C.2. The test substance was completely soluble in test medium at the concentrations tested. The entire project was performed applying semi-static test conditions, with renewal of test solutions after 24 hours of exposure, because the test substance was known to be highly unstable in water. Test solutions were prepared starting with a stock solution at 100 mg/L. The initial test solution was clear and colourless but, almost instantaneous after dissolving the test substance in the test medium, the solution turned increasingly hazy and the pH dropped to acidic levels. The pH was adjusted to approximately 8.5 as a consequence of the observed steep pH fall.

The project started with a range-finding test exposing ten daphnia per concentration (in duplicate, 5 per vessel) to a blank control and a test substance filtrate prepared at a loading rate of 100 mg/L and dilutions of the filtrate containing 0.1, 1.0 and 10 % of the filtrate. No immobile daphnia were observed during the 48-hour exposure period in the undiluted filtrate or any of the filtrate dilutions tested. Hence, the expected 48h EC50 exceeded the concentration present in a filtered solution prepared at a loading rate of 100 mg/L.

The project was continued with a limit test exposing twenty daphnia per concentration (in quadruplicate, 5 per vessel) to a blank control and a filtered test substance solution prepared at a loading rate of 100 mg/L. The total test period was 48 hours. Samples for analytical confirmation of actual exposure concentrations were taken from the freshly prepared solutions at the start and after 24 hours of exposure and at the end of the test period.

Analysis of the samples taken during the limit test showed that measured lithium concentrations were stable between 3.4 and 3.6 mg/L during the 24-hour refreshment periods and in agreement with what was expected based on the theoretical percentage of lithium (3.58 %) in the test substance. All results will however be based on the loading rate as measurement of lithium can only be considered indicative for the real test substance concentration present in the test solution.

The study met the acceptability criteria prescribed by the protocol and was considered valid.

The test substance did not induce acute immobilisation of Daphnia magna at a loading rate of 100 mg/L after 48 hours of exposure (NOEC).

The 48h EC50 exceeded a loading rate of 100 mg/L, the regulatory limit concentration. (NOTOX, 2004)

Long-term toxicity to aquatic invertebrates

The chronic toxicity of Lithium-bis(oxalate) borate (LIBOB) towards Daphnia magna has been investigated in an 21 -day Reproduction Test according to OECD Guideline 211 and in compliance with GLP criteria. The test item was not completely soluble in test medium at the initial prepared concentration of 100 mg/L and therefore the hazy dispersions were filtered through a rough paper filter to remove any precipitate. The resulting Saturated Solutions (SS) were clear and colourless, and used as the highest test concentration. Lower test concentrations were prepared by subsequent dilutions of the SS. All final test solutions were clear and colourless.

The reproduction test was performed in a semi-static system and comprised of 10 vessels per test concentration and 20 vessels for an untreated control group. Each of the vessels contained one neonate (<24h old) Daphnia magna in 50 ml test medium. The Lithium-bis(oxalate) borate (LiBOB) groups tested contained 1.0, 3.2, 10, 32 and 100% of an SS prepared at a loading rate of 100 mg/L. The study duration was 21 days and the test solutions were renewed every 48 hours. The daphnids were fed on a daily basis with a Chlorella pyrenoidosa/Marinure suspension. Every workday, the condition of the parental daphnids was recorded, and during the reproduction phase, the number of living offspring, immobile young and appearance of unhatched (aborted) eggs was recorded. At the end of the test, the lengths of the surviving parental daphnids were measured.

During the study, samples for chemical analyses were taken at the beginning and the end of three intervals of 48 hours. Analyses were based on Lithium concentrations and the measured concentrations were in agreement with concentrations expected based on the theoretical fraction of Lithium (3.57%), the applied loading rate and the respective dilution factor. The measured Lithium concentrations remained stable during the 48-hour renewal intervals. Results were based on both the loading rate, and percentages thereof, and based on the expected concentration in solution (based on Li measurements). Measurement of Lithium is indicative of the remaining Lithium-bis(oxalate)borate concentration present in the test solutions.

No statistically significant mortality was observed at any of the concentrations tested.

The onset of reproduction was significantly affected at the highest concentration tested.

The number of produced offspring and the growth of the parental daphnids were significantly affected at the two highest concentrations tested.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

The 21 -day NOEC for reproduction has been determined to be 10 mg/L nominal and the EC₁₀ for reproduction has been determined to be in the range of 19-43 mg/L nominal (19 mg/L nominal), respectively.

The EC₅₀ for reproduction was > 100 mg/L nominal, NOEC mortality was 10 mg/L and NOEC growth was 10 mg/L. (WIL, 2016)

Toxicity to aquatic algae and cyanobacteria

The present study examined the influence of the test item lithium bis(oxalato)borate (LiBOB) on the growth of an unicellular green algae- Pseudokirchneriella subcapitata- according to OECD guideline 201 and EU method C.3. The test concentrations (nominal 5.0 to 50.0 mg/L) were selected according to results of a range-finding test. The selected test concentrations were analysed at the beginning and at the end of the study, ensuring that the nominal values could be used for calculations. During the test time, the control algae cells growth was as expected and pH values and temperature were steady.  After exposure for 72 hours, the test item, in the concentration range of 20.0 to 50.0 mg, had a statistically significant inhibitory effect on the growth based on the average specific growth rate, yield and areas under the growth curves of Pseudokirchneriella subcapitata. The 72-h-EC50 parameters (EbC50, Er50, EyC50) are detailed as follows: 35.11 ml/L (biomass), 40.69 mg/L (growth rate), 34.24 mg/L (yield). The overall NOEC was determined to be 10.0 mg/L. Additionally, no abnormalities of the algae cells were observed. (LAB, 2008)

Toxicity to microorganisms

1. key study

The purpose of the 3 hour test was to evaluate the influence of the test item lithium bis(oxalato)borate (LiBOB) on the activity of activated sludge by measuring the respiration rate under defined conditions. The respiration rates (total, heterotrophic and nitrification oxygen uptake rates) of samples of activated sludge fed with synthetic sewage were measured in an enclosed cell containing an oxygen electrode after a contact time of 3 hours. The test item affected the pH within the test system, therefore the test item treatments were investigated in duplicate series: with and without pH adjustment. Based on the preliminary information about the test item, lithium bis(oxalato)borate (LiBOB) was investigated at the nominal concentrations of 62.5; 100; 160; 256 and 410 mg/L without pH adjustment and at the concentrations of 100; 160; 256; 410 and 655 mg/L with pH adjustment. Defined amounts of the test item were weighed directly into the test vessels in both cases. 2, 7 and 24.5 mg of 3,5-Dichlorophenol /L served as positive reference control. Further blank (inoculum) controls and nitrification controls were examined. The main test was performed without abiotic controls, based on the results of the preliminary test where abiotic controls were tested at the test item concentration of 1000 mg/L and no remarkable abiotic oxygen consumption was noticed. All validity criteria of the study were met. Based on measured inhibition rates the 3 hour EC10, EC50 and EC80 and their 95 %-confidence limits were calculated by Probit analysis using TOXSTAT software.

The results without pH adjustment:

EC10: 122.37 mg/L (109.25–137.06)

EC50: 228.69 mg/L (212.73–245.85)

EC80: 344.83 mg/L (311.54–381.68)

The EC50 value was determined as 228.69 mg/L, consequently: EC50 > 100 mg/L.

The results with pH adjustment:

EC10: 211.29 mg/L (187.86–237.64)

EC50: 421.62 mg/L (388.11–458.03)

EC80: 663.70 mg/L (584.13–754.09)

The EC50 value was determined as 421.62 mg/L, consequently: EC50 > 100 mg/L. The resulting overall NOEC is 100 mg/L. (Toxicoop, 2012)

2. disregarded study

Lithium bis(oxalato)borate (LiBOB) was tested for toxicity to microorganisms by determination of inhibition of the respiration of activated sludge similar to OECD guideline 209. Three experiments were performed. In the pre-tests, LiBOB was tested using three concentrations (10, 100, 1000 mg/L) for range-finding and 1000 mg/L for the limit test. Duration of the tests was 30 min and 3 hours. The inoculum was taken from a domestic sewage treatment plant and was washed before use. The concentration of activated sludge in the test medium was 1.5 g/L. The EC50 (3 h) of 3,5-Dichlorophenol was 22.68 mg/L, which lies in the recommended range of 2 mg/L to 25 mg/L for total respiration. An inhibition of the respiration was detected in both the range-finding and the limit test. The main test was performed with the following concentrations: 25.6, 64, 180, 400, 1000 mg/L. Duration of the test was 30 min. The following result for LiBOB was determined: 30 min EC50 = 195.06 mg/L. (APM, 2011)

Discussion

The study report of APM is exhibiting a few insufficiencies with regards to the used OECD guideline 209, leading to the categorisation of Klimish 3. The OECD guideline recommends six controls and five treatment concentrations in a geometric series with five replicates in order to obtain both a NOEC and an EC. Only one control group and no replicates had been used in order to validate the observed results. The duration of treatment was three hours and/or 30 min, respectively, in the performed range-finding- as well as in the limit-test. In the definitive test only the 30 min exposure time was tested without any reasonable explanation. The OECD guideline further states that when a substance adversely affects the pH value of the treatment solution a pH adjustment should be implemented in the study record in order to assess the effect of the test substance on the sludge with and without pH adjustment. This was not done in this study although there was a change of pH in the definitive test of up to 20 % after 30 min of exposure. The report of the results in this study was very poor as there were oversights regarding the test substance final concentration in the test suspension of the activated sludge. Instead of doubling the used amount of test substance in order to get the nominal concentration, it was halved (according to the table under 13.2). Further in the raw data the concentrations of the range finding test was registered in the table of the definitive test (see table 19.5). Taking these insufficiencies into account the results of the TOXICOOP study from 2012 are used for the chemical safety assessment.