Disodium piperazine-1,4-diethanesulphonate

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:

23 Oct 2017 to 21 Jan 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Deviations:

yes

Remarks:

None of the deviations were considered to have impacted the overall integrity of the study or the interpretation of the study results and conclusions.

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test item information
Appearance: White powder
Test item storage: At room temperature
Stable under storage conditions until: 30 November 2018 (retest date)

Analytical monitoring:

yes

Details on sampling:

Samples for possible analysis were taken from all test concentrations and the control according to the schedule below.

Frequency: at t=0 h, t=24 h and t=72 h
Volume: 2.0 mL
Storage: Samples were stored in a freezer (≤-15°C) until analysis at the analytical laboratory of the Test Facility.

At the end of the exposure period, the replicates with algae were pooled at each concentration before sampling.

Compliance with the quality criteria regarding maintenance of actual concentrations was checked by running a test vessel at the highest test item concentration but without algae and samples for analysis were taken at the start, after 24 hours of exposure and at the end of the test period.

Additionally, reserve samples of 2.0 mL were taken from all test solutions for possible analysis. If not already used, these samples were stored in a freezer (≤-15°C) for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.

Vehicle:

not specified

Details on test solutions:

The batch of 1,4 piperazinediethanesulfonic acid disodium salt tested was a white powder of high purity and completely soluble in test medium at the concentrations tested. No correction was made for the purity/composition of the test item. Preparation of test solutions started with the highest concentration of 100 mg/L. No other treatment than vigorous shaking was needed to completely dissolve the test item in test medium. Lower test concentrations were prepared by subsequent dilutions of the highest concentration in test medium. All test solutions were clear and colorless at the end of the preparation procedure. After preparation, volumes of 50 mL were added to each replicate of the respective test concentration. Subsequently, 1 mL of an algal suspension was added to each replicate providing a cell density of 10 to the power of 4 cells/mL. Due to the observed degree of adsorption during the analytical method development, all glass ware employed in this study was silanized prior to use. Any residual volumes were discarded.

Test organisms (species):

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

Details on test organisms:

Species: Pseudokirchneriella subcapitata, strain: NIVA CHL 1
Source: In-house laboratory culture.
Reason for selection: This system is a unicellular algal species sensitive to toxic items in the aquatic ecosystem and has been selected as an internationally accepted species.

Fresh Water Algae Culture
Stock culture: Algae stock cultures were started by inoculating growth medium with algal cells from a pure culture on agar. The suspensions were continuously aerated and exposed to light in a climate room at a temperature of 21-24°C.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Hardness:

Hardness (Ca+Mg) 0.24 mmol/L (24 mg CaCO/L)

Test temperature:

During the exposure period the temperature measured in the incubator was maintained between 23 and 24°C. Temperature remained within the limits prescribed by the study plan (21-24°C, constant within 2°C).

pH:

pH: 8.1 ± 0.2

Nominal and measured concentrations:

Six exponentially growing algal cultures per group were exposed to an untreated control and to a nominal concentration of 100 mg/L in the limit test

Details on test conditions:

Test duration: 72 hours
Test type: Static
Test vessels: 100 mL, all-glass, silanized, containing 50 mL of test solution
Medium: M2
Cell density: An initial cell density of 1 x 10 to the power of 4 cells/mL.
Illumination: Continuously using TLD-lamps2
Incubation: Capped vessels were distributed at random in the incubator and daily repositioned. During incubation the algal cells were kept in suspension by continuous shaking.

Reference substance (positive control):

not specified

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Key result

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 70 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Yield

Details on results:

Results are attached in the background information

Reported statistics and error estimates:

For determination of the NOEC and the EC50 the approaches recommended in the OECD guideline 201 were used. An effect was considered to be significant if statistical analysis of the data obtained for the test concentrations compared with those obtained in the negative control revealed significant inhibition of growth rate or inhibition of yield (Two-sample t-test Procedure, α=0.05, one-sided, smaller). Additionally, the EC10 and EC20 were determined to meet the recommendations as put down in "A Review of Statistical Data Analysis and Experimental Design in OECD Aquatic Toxicology Test Guidelines" by S. Pack, August 1993. No EC50-values could be calculated because the test item proved to be non-toxic (EC50 > maximum soluble concentration tested). The calculations were performed with ToxRat Professional v. 3.2.1. (ToxRat Solutions GmbH, Germany).

Validity criteria fulfilled:

yes

Conclusions:

In conclusion, under the conditions of the present study with Pseudokirchneriella subcapitata, no inhibition of growth rate or inhibition of yield was recorded at any of the concentrations of 1,4 piperazinediethanesulfonic acid disodium salt tested. The EC50 for growth rate inhibition (72h-ERC50) was beyond the range tested, i.e. exceeded a measured concentration of 70 mg/L being considered to be the maximum concentration soluble in test medium at a loading rate of 100 mg/L. The EC50 for yield inhibition (72h-EYC50) was beyond the range tested, i.e. exceeded a measured concentration of 70 mg/L being considered to be the maximum concentration soluble in test medium at a loading rate of 100 mg/L. The 72h-NOEC for growth rate inhibition and the 72h-NOEC for yield inhibition were 70 mg/L.

Executive summary:

The objective of the study was to evaluate 1,4 piperazinediethanesulfonic acid disodium salt for its ability to generate toxic effects in Pseudokirchneriella subcapitata during an exposure period of 72 hours and, if possible, to determine the NOEC, EC10 and EC50 for both inhibition of growth rate and inhibition of yield. The study procedures described in this report were based on the OECD guideline No. 201, 2006; Annex 5 corrected 28 July 2011.  

The batch of 1,4 piperazinediethanesulfonic acid disodium salt tested was a white powder of high purity and completely soluble in test medium at the concentrations tested.  To prevent adsorption, all glass ware employed in this study was silanized prior to use. A combined limit/range-finding test was performed. Six exponentially growing algal cultures per group were exposed to an untreated control and to a nominal concentration of 100 mg/L in the limit test. In addition, three replicates per group were exposed to 0.10, 1.0 and 10 mg 1,4 piperazinediethanesulfonic acid disodium salt per litre in a combined range-finding test. The initial algal cell density was 10 to the power of 4 cells/mL. The total exposure period was 72 hours and samples for analytical confirmation of actual exposure concentrations were taken at the start, after 24 and 72 hours of exposure.  No inhibition of growth rate or yield was recorded at any of the concentrations tested when compared to the control group.  Samples taken from the limit concentration were analysed. At the start of the test, the actual test concentrations were 70 and 71 mg/L in the samples with and without algae, respectively. The measured concentrations remained stable throughout the test duration, i.e. were at 96-107% of the initial concentrations at the end of the test. Based on these results, the measured concentrations were used to determine the effect parameters. The study met the acceptability criteria prescribed by the study plan and was considered

valid. The effect parameters obtained in this study are summarized below.

Parameter (mg/L)              NOEC              EC10              EC20              EC50

Growth rate                          70                  >70 >70 >70

Yield 70 >70 >70 >70

In conclusion, under the conditions of the present study with Pseudokirchneriella subcapitata, no inhibition of growth rate or inhibition of yield was recorded at any of the concentrations of 1,4 piperazinediethanesulfonic acid disodium salt tested.

 

Additional information

In conclusion, under the conditions of the present study with Pseudokirchneriella subcapitata, no inhibition of growth rate or inhibition of yield was recorded at any of the concentrations of 1,4 piperazinediethanesulfonic acid disodium salt tested. The EC50 for growth rate inhibition (72h-ERC50) was beyond the range tested, i.e. exceeded a measured concentration of 70 mg/L being considered to be the maximum concentration soluble in test medium at a loading rate of 100 mg/L. The EC50 for yield inhibition (72h-EYC50) was beyond the range tested, i.e. exceeded a measured concentration of 70 mg/L being considered to be the maximum concentration soluble in test medium at a loading rate of 100 mg/L. The 72h-NOEC for growth rate inhibition and the 72h-NOEC for yield inhibition were 70 mg/L.