Calcium cyanamide

Additional ecotoxological information

Administrative data

Endpoint:

additional ecotoxicological information

Remarks:

Outdoor mesocosm study

Type of information:

experimental study

Adequacy of study:

key study

Study period:

June 13, 2018: single application was conducted. Fate and effects were monitored over approximately twelve weeks after the application.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Report date:

2019

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

other: OECD, 2006. Guidance Document on Simulated Freshwater Lentic Field Tests (Outdoor Microcosms and Mesocosms). OECD Environment Health and Safety Publications Series on Testing and Assessment No. 53. Paris.

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EFSA PPR Panel (EFSA Panel on Plant Protection Products and their Residues), 2013. Guidance on tiered risk assessment for plant protection products for aquatic organisms in edge-of-field surface waters. EFSA Journal 2013, 11(7): 3290, 268 pp.

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: European Chemicals Agency, 2017. Guidance on Biocidal Products Regulation: Volume IV Environment - Assessment and Evaluation (Parts B+C). Version 2.0 October 2017. Helsinki.

GLP compliance:

yes (incl. QA statement)

Remarks:

A GLP outdoor mesocosm study was performed at the Fraunhofer Institute for Molecular Biology and Applied Ecology (IME) in cooperation with Mesocosm GmbH.

Type of study / information:

Outdoor mesocosm study: The aim of this study is to investigate effects of cyanamide, the first and essential metabolite of the mineral fertiliser PERLKA, mainly consisting of calcium cyanamide, on freshwater ecosystems by monitoring zooplankton, macroinvertebrates, phytoplankton, periphyton and macrophytes in lentic outdoor mesocosms (enclosure systems). Fate and effects of the test item were monitored over approximately twelve weeks after the application.

Test material

Test material information

Constituent 1

Reference substance name:

Cyanamide

EC Number:

206-992-3

EC Name:

Cyanamide

Cas Number:

420-04-2

Molecular formula:

CH2N2

IUPAC Name:

cyanamide

impurity 1

Reference substance name:

Cyanoguanidine

EC Number:

207-312-8

EC Name:

Cyanoguanidine

Cas Number:

461-58-5

Molecular formula:

C2H4N4

IUPAC Name:

2-cyanoguanidine

impurity 2

Reference substance name:

Water

EC Number:

231-791-2

EC Name:

Water

Cas Number:

7732-18-5

Molecular formula:

H2O

IUPAC Name:

Dihydrogen oxide

impurity 3

Reference substance name:

Urea

EC Number:

200-315-5

EC Name:

Urea

Cas Number:

57-13-6

Molecular formula:

CH4N2O

IUPAC Name:

urea

additive 1

Reference substance name:

Reaction mass of 1,2-propanediol, 1,2-diformate and 1,2-propanediol, 1-formate and 1,2-propanediol, 2-formate and formic acid and propane-1,2-diol

EC Number:

940-273-1

Molecular formula:

not applicable (multi-cont. substance)

IUPAC Name:

Reaction mass of 1,2-propanediol, 1,2-diformate and 1,2-propanediol, 1-formate and 1,2-propanediol, 2-formate and formic acid and propane-1,2-diol

Test material form:

solid: crystalline

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: 806501
- Expiration date of the lot/batch: 2019-03-06

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Store between < 35°C, keep away from direct sunlight. Materials to avoid: Acids and bases, Flammable materials
- Stability under test conditions: No decomposition if stored and applied as directed. Violent, exothermic reaction with acids, bases and temperatures above 40°C.

OTHER SPECIFICS:
- Water solubility: 850 g/L (at 25°C)
- Vapour pressure: 0.005 hPa (at 20°C)
- Density: 1.28 g/cm³ (at 20°C)

Results and discussion

Any other information on results incl. tables

One day after application, the recovery of the nominal concentrations of cyanamide in the enclosure water was 97 % on average and ranged between about 84 and 110 %. Thus, the intended initial exposure was confirmed, and nominal test concentrations were used to characterize the exposure. The variability of the measured cyanamide concentrations between the replicates per test concentration was low and the general pattern of dissipation was similar over the different test concentrations. However, DT50 values varied between 3 and 28 days and increased with nominal concentration, providing an overall mean of 13 days.

A large variety of species covering several relevant taxonomic groups of freshwater ecosystems were present in the test systems, representing zooplankton, macroinvertebrates (incl. crustaceans, insects, snails and oligochaetes), algae and macrophytes. For eight macroinvertebrate, nine zooplankton, nine phytoplankton and five macrophyte taxa, the calculated Minimum Detectable Differences (MDDs) were sufficiently low to allow a reliable analysis according to the proposal of Brock et al. (2015). Thus, the requirements of the EFSA aquatic guidance document (EFSA 2013) that at least eight potentially sensitive populations should allow a statistical evaluation of direct effects are clearly fulfilled.

Direct effects of Cyanamide were found on crustaceans, insects and algae of the class Cryptophyceae. Indirect effects, i.e. increased abundances compared to controls, were observed for rotifers, Tubificidae and several classes of the phytoplankton, periphyton and Lemna.

The following effects were found in the different test concentrations (see also Table 1):

 

0.032 mg/L

Almost all taxa and endpoints were not affected. Only the Cladocera Alona sp. of the zooplankton and the Cryptophyceae Chroomonas acuta / Rhodomonas sp.in the phytoplankton might have been slightly and temporarily affected (effect class 2). However, abundance of Alona sp. was generally low and the NOECs < 0.032 mg/L were found on the only dates when the species was not absent in at least one control ample. No clear decline in abundance of Alona sp. was observed up to 0.32 mg/L. The reduction of Chroomonas acuta in 0.032 mg/L was restricted to day 6 and the mean was still within the range of the control.

 

0.1 mg/L

Additional slight effects were found for Cyclopidae, Chaoborus larvae, and Chlorophyceae. Effects on the community structure of the zooplankton were considered pronounced but temporary (class 3A) since significant deviations were found on two consecutive samplings (day 30 and 44). However, no pronounced effects on populations were found. Effects on the macroinvertebrate community structure were slight (due to the slight effects on Chaoborus).

 

0.32 mg/L

Additional taxa showed slight effects and pronounced temporary direct or indirect effects (class 3A) were found for more taxa, e.g. Alona sp., Chaoborus sp. and Chroomonas acuta, Chlorophyceae). Only the PRCs for the phytoplankton indicated a long-term effect on the community structure (class 5B).

 

1.0 mg/L

Some zooplankton taxa showed pronounced effects, some of them at the end of the study which did not allow to assess the duration of these effects. From the macroinvertebrates, the damselflies (Zygoptera) might have been slightly affected but Asellus aquaticus was affected over more than eight weeks but recovered until the end of the study (class 5A). Effects on the macroinvertebrate community structure were considered long-term without full recovery (class 5B).

 

3.2 mg/L

Several taxa showed long-term effects, often until the end of the study. Crustaceans, insects and Cryptophyceae were directly affected while rotifers and the total phytoplankton were promoted until the end of the study. A temporary promotion was also found for Lemna sp. Other macrophytes as well as snails were not affected.

 

The intended exposure to cyanamide was confirmed and the dissipation over time was characterized. Effects could be statistically analysed for a variety of populations covering several major taxonomic groups and also for the zooplankton, macroinvertebrate and phytoplankton community. From the effect classification (Table 1) based on the statistical analysis and the interpretation of the course of abundances or other endpoints over time, the following effect concentrations are proposed to derive an ETO and an ERO-RAC:

At 0.1 mg/L, no pronounced effects (class 3A or higher) on populations were found, only the effect on the zooplankton community structure was considered class 3A for two sampling dates. Short-term effects on the zooplankton community are not considered relevant and therefore 0.1 mg/L de facto represents a Class 2 concentration. Thus, this concentration would still allow an estimation of a protective ETO-RAC for all populations tested.

At 0.32 mg/L, pronounced effects on a few populations were found, but these were only temporary (class 3A). However, indicated by the ordination analysis, the phytoplankton community structure was affected until the end of the study, which might have been a result of the high dynamic of the phytoplankton and promotion of some species, especially green algae at the end of the study which were still rare compared to the total phytoplankton. Thus, the ecological relevance of these promotions of a few, not dominant algae species is probably small and the ERO-RAC could thus be derived also from this concentration considering that no population showed long-term effects.

Table 1. Effect classification. For definition of classes see EFSA PPR 2013 and Brock et al. 2015:

1 = no effect, 2 = slight effect, 3A = effect duration shorter than 8 weeks, 4A = effect at the end of the study, duration could not be assessed,

5A = effect over more than 8 weeks but recovery within the study, 5B = effect longer than 8 weeks without recovery within the study.

	Nominal test concentration
	0.032 mg/L	0.1 mg/L	0.32 mg/L	1 mg/L	3.2 mg/L
Asellus aquaticus	1	1	1	5B	5B
Chaoborus sp.	1	2	3A	3A	5B
Cloeon dipterum	1	1	1	2	5A
Lymaeidae	1	1	1	1	1
Planorbidae	1	1	1	1	1
Tubificidae	1	1	1	1	2+
Zygoptera	1	1	1	3A-4A	3A - 4A
Lemna gibba	1	1	1	3A+	3A+
Mentha aquatica	1	1	1	1	2
Myriophyllum spicatum	1	1	1	1	2+
Other macrophytes	1	1	1	1	1
Total phytoplankton	1	1	1	1	3A+ - 5B+
Chroomonas sp.	2	2	(5A)	3A	5A - 5B
Cryptomonas sp.	1	1	1	3A	5B
Nitzschia palea	2+	2+	3A+ - 5B+	5A+ - 5B+	5A+ - 5B+
Total phytopl. Chlo a	1	1	1	2/2+	3A / 5B+
Crytophyceae Chlo a	1	1	2	3A	5B
Periphyton Chl a	1	1	1	1	3A+
Primary production	1	1	1	1	1
Total assessment	2	2	3 / 3+	5B	5B

Based on this evaluation, an ETO-RAC (regulatory acceptable concentration using the ecological threshold option) could be derived based on the 0.1 mg/L test concentration since no or only slight effects were found here. 

For the Ecological Recovery Option, conservatively also the 0.1 mg/L would be used. However, since forChroomonas sp. recovery was demonstrated in the enclosures treated with 1 mg/L the missing recovery at 0.32 mg/L was probably not caused by the treatment and a substance related promotion of the cyanobacteriumNitzschia paleaat 0.32 mg/L was not clearly persistent over several weeks, the ERO-RAC might instead be derived from the 0.32 mg/L test concentration.

Applicant's summary and conclusion

Conclusions:

The derivation of a regulatory acceptable concentration (RAC) for cyanamide is based on two options: (1) the ecological threshold option (ETO), accepting negligible population effects only, and (2) the ecological recovery option (ERO), accepting some population-level effects if ecological recovery takes place within an acceptable time period. Based on this evaluation, an ETO-RAC (regulatory acceptable concentration using the ecological threshold option) could be derived based on the 0.1 mg/L test concentration since no or only slight effects were found here. Based on the preliminary evaluation of the mesocosm study, acceptable short-term effects (effect class 3A, EFSA PPR 2013) were found at 0.32 mg/L (= ERO-RAC).

Executive summary:

This mesocosm study was conducted in accordance with the OECD Guidance Document “Freshwater Lentic Field Tests” (2013) and the recommendations from the EFSA PPR Panel (2013) and the Biocide guidance (2017). Observed effects of the test item are classified according to the EFSA (2013) and Brock et al. (2015). The aim of this study is to investigate effects of cyanamide, the first and essential metabolite of a calcium cynamide containing mineral fertilizer, on freshwater ecosystems by monitoring zooplankton, macroinvertebrates, phytoplankton, periphyton and macrophytes in lentic outdoor mesocosms (enclosure systems). The mesocosms contained an indigenous assemblage of invertebrates (zooplankton, macroinvertebrates) and plants (phytoplankton, periphyton and macrophytes). In addition, three macrophyte species (Lemna sp., Myriophyllum spicatum and Mentha aquatica) were introduced in pots or swimming enclosures to monitor effects on growth. The tests was conducted using 15 enclosures set up in an artificial pond at the test site. The pond was set up with sediment and water from a nearby pond in October 2016 allowing the establishment of a natural community of phytoplankton, periphyton, macrophytes, zooplankton and macroinvertebrates. The stainless steel enclosures were introduced on May 11, 2018. With an average depth of the water body of e.g. 110 cm the total water volume per enclosure was approximately 1800 L. Five of the enclosures served as untreated controls while ten enclosures (two replicates per concentration) were treated with five concentrations of the test item cyanamide: 0.032; 0.1; 0.32; 1.0; and 3.2 mg/L. The single application was conducted on June 13, 2018 by using separation funnels to distribute the application solutions directly in to the water column.

The mean recoveries of the nominal concentrations of cyanamide in the enclosure water in the first three samplings, 1, 3 hours and 1 day after the application, were 80 %, 90 % and 97 %, respectively. Thus, the intended dosing of the test systems was confirmed and the exposure levels are expressed as nominal concentrations.

The variability of measured concentrations between the replicates per test concentration was very low at concentrations above 0.1 mg/L and the general pattern over time was similar over the different test concentrations. However, dissipation seems to have been slower at higher test concentrations. For a conservative estimation, the DT50 was calculated based on the data of the highest test concentration only by log-linear regression and found to be 27 d.

Based on the MDDs for the evaluated data, the criterion by EFSA PPR (2013) that a statistical analysis of direct effects should be possible for at least eight potentially sensitive taxa is fulfilled since seven macroinvertebrate taxa (Asellus aquaticus, Chaoborus sp., Cloeon dipterum, Lymnaea stagnalis, Planorbidae, Tubificidae, Zygoptera), seven phytoplankton taxa (Ankyra judayi, Characium ensiforme, Chromulina minima cf, Chroomonas acuta/ Rhodomonas sp, Chrysophyceae (1-5-µm), Chrysophyceae (6-10 µm), Cryptomonas erosa + ovata, Desmarella moniliformis) and five macrophyte species (Lemna gibba, Mentha aquatica, Myriophyllum spicatum, Chara globularis, Potamogeton natans) fulfil the criterion proposed by Brock et al. (2015).

 

Direct effects of cyanamide were found on algae and macroinvertebrates. Algae of the class Cryptophyceae were the most sensitive group found in the study with slight temporary effects at 0.032 and 0.1 mg/L on Chroomonas sp. and pronounced effects at 0.32 mg/L and higher. Recovery was clearly demonstrated at 1 mg/L and thus, the more pro-longed deviations from controls at 0.32 mg/L are probably not treatment related. At 3.2 mg/L no recovery was observed.

For another species, Cryptomonas sp., no effects were found up to 0.32 mg/L but pronounced effects with and without recovery were found at 1 and 3.2 mg/L, respectively. Other algae, e.g. the cyanobacterium Nitzschia palea,were promoted. These indirect effects were very slight at 0.032 and 0.1 mg/L.

Effects on chlorophyll a concentrations of phytoplankton and periphyton were less pronounced as for the cell abundances of phytoplankton species. Also the macrophytes were less affected. As slight growth inhibition was only found for Mentha aquatica at 3.2 mg/L. The duck weed Lemna gibba, was temporarily promoted at 1 and 3.2 mg/L. Total photosynthesis indicated by dissolved oxygen concentration, pH and conductivity of the enclosure water was not affected.

 

From the macroinvertebrates, the midge Chaoborus sp. was found to be the most sensitive species with slight effects found at 0.1 mg/L pronounced effects with recovery within eight weeks up to 1 mg/L, but no recovery within the study at 3.2 mg/L (Figure4). AlsoAsellus aquaticus(Isopoda) and Cloeon dipterum (Ephemeroptera) were affected over more than eight weeks at the highest test concentrations, but showed no effects up to 0.32 mg/L. Full recovery of Asellus in the enclosures treated with 1 mg/L could not be demonstrated since mean abundance at the end of the test was still less than 50 % of the mean control abundance.

Based on this evaluation, an ETO-RAC could be derived based on the effect class 2 concentration of 0.1 mg/L test concentration since no or only slight effects were found here.

Since only short-term exposure from the use of PERLKA is predicted by the exposure modelling for edge-of-field water bodies, also ecological recovery can be taken into account. Based on the preliminary evaluation of the mesocosm study, acceptable short-term effects (effect class 3A, EFSA PPR 2013) were found at 0.32 mg/L (= ERO-RAC).