Flue dust, portland cement

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-08-17 to 2010-09-09

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test organisms (species):

Daphnia magna

Details on test organisms:

Daphnia magna STRAUS (Clone 5)

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

21 d

Hardness:

Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L

Test temperature:

20+/-2 °C

pH:

Additionally 10 replicates of the highest tested WAF (100 mg/L) adjusted to a pH in the range 7.0  0.2 by addition of 1 M HCl

Nominal and measured concentrations:

The following water accommodated fractions (WAF) were tested with a dilution factor of 2: 6.25 - 12.5 - 25 - 50 and 100 mg/L. The loading levels are based on the results of an acute immobilization test performed at the test facility (Noack, M. Flue Dust T Acute Immobilization Test to Daphnia magna, Static, 48 h (NOACK Lab-ID: DAI13593/091218EL), DR. U. NOACK-LABORATORIEN, Sarstedt, Germany, 2010)

Details on test conditions:

Water accommodated fractions (WAF) of 6.25 - 12.5 - 25 - 50 - 100 mg/L were prepared with dilution water one day prior to application and at every water renewal. The WAF’s were shaken with 20 rpm for 24 h at room temperature. After a separation phase of at least 1 h the WAF’s were taken from the homogeneous liquid phase.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

21 d

Dose descriptor:

EL10

Effect conc.:

ca. 68.2 mg/L

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

reproduction

Reproductive output

The number of juveniles in all replicates of control, the pH-control and all test groups was recorded every day. Results concerning the number of juveniles of all test and control groups for every test day are presented inTable 10toTable16.

The number of juveniles produced in the 10 replicates of the test and control groups between the 8th and 21st day are presented inTable3.

Four broods were released by all surviving animals of the control, the pH-control and the loading levels of 6.25 to 50.0mg/L. In the loading level 100 mg/L four of nine daphnids had less than four broods with living juveniles. There were one parental daphnid having only one brood and three parental daphnids having three broods.

The average number of juveniles per parent alive at the end of the test in the control group was 90.0 after 21 days.The reproductive outputwas statistically significant reduced in the nominal loading level of 100 mg/L when compared to the control (One Way Analysis of Variance,Dunnett’smethod, p = 0.05).The EL₁₀-value for the reduction of the reproductive output was calculated by sigmoidal dose-response regression to be 68.2 mg/L. An EL₅₀-value for the reproductive output was not determinable because no effects≥50 % (reduction of the reproductive output) occurred within the tested concentration range (Figure1).

The coefficient of variation of the number of living offspringproduced per parent was 9 % in the control. In the loading levels of 6.25 to 50.0 mg/L and the pH-control the calculated coefficients of variation were in the range of 10 to 13 % and comparable to the control. In the loading level of 100 mg/L the coefficient of variation increased to 22 %.

 

Table3:   Number of Juveniles in the Control and Test Groups after 21 Days

Nominal Loading Level Of the Test Item   [mg/L]												No. of Parents prod. Juv.	Mean No. of Juveniles per Parent producing Juveniles			Comparison versus Control
	Number of Juveniles in Replicate No.										Total No.				CV
																Red.	Stat.
	1	2	3	4	5	6	7	8	9	10	å	N	MV±SD		[%]	[%]
100 (pH-control)*	97	78	106	103	103	101	104	81	98	101	972	10	97±	10	10	-8	no
100	58	58	64	65	--	59	83	83	50	40	560	9	62±	14	22	38	yes
50.0	106	107	88	86	99	110	99	113	106	88	1002	10	100±	10	10	-11	no
25.0	94	93	85	102	102	103	117	99	101	82	978	10	98±	10	10	-9	no
12.5	58	90	82	98	83	100	85	87	90	89	862	10	86±	12	13	4	no
6.25	105	87	85	92	84	106	100	89	93	111	952	10	95±	10	10	-6	no
Control	94	99	106	87	85	88	84	93	84	80	900	10	90±	8	9	—	—

N                         = Number of parental daphnids producing juveniles alive

MV ± SD             = Mean value ± standard deviation

CV                      = Coefficient of variation

Red.                   = Reduction of the reproductive output (negative values mean increase of the reproductive output)

Stat.                    =Statistically significant difference (One Way Analysis of Variance, Dunnett’smethod, p = 0.05)

yes                    =Statistically significant

no                      = Statistically not significant

--                         = Not applicable, due to mortality of the parental daphnid

*                          = Adjusted with 1 M NaOH to a pH-value of 7 ± 0.2

Intrinsic Rates of Natural Increase (IR)

Theintrinsic rates of natural increase (IR)of the surviving parent animals accounting for generation time and number of offspring were used for calculation of population growth and maintenance. The mean IR of the surviving daphnids of the treatment groups were compared to the control by Kruskal-Wallis One Way Analysis of Variance on Ranks,Dunn’smethod (p = 0.05). There was a statistical significance in the tested loading level of 100 mg/L when compared to the control.The calculated IR of the control and test groups are presented in Table 4, mean values are presented in Figure 2.

Table4:       Intrinsic Rates of Natural Increase

Nominal Loading Level of the Test Item [mg/L]	Intrinsic Rate of Natural Increase in Replicate No.										Mean IR		CV
	1	2	3	4	5	6	7	8	9	10	MV±SD		[%]
100 (pH-control)*	0.48	0.49	0.48	0.48	0.47	0.51	0.47	0.47	0.49	0.47	0.48	±0.01	3
100	0.38	0.39	0.39	0.47	--	0.40	0.42	0.51	0.37	0.30	0.40	±0.06^1)	15
50.0	0.52	0.52	0.49	0.45	0.50	0.51	0.50	0.52	0.49	0.50	0.50	±0.02	4
25.0	0.49	0.48	0.46	0.51	0.50	0.47	0.50	0.50	0.49	0.49	0.49	±0.01	3
12.5	0.47	0.49	0.50	0.50	0.51	0.51	0.49	0.49	0.51	0.49	0.49	±0.01	3
6.25	0.52	0.49	0.48	0.47	0.48	0.48	0.50	0.50	0.49	0.53	0.49	±0.02	3
Control	0.51	0.50	0.52	0.50	0.47	0.50	0.49	0.50	0.49	0.48	0.50	±0.01	3

IR               = Intrinsic rate of natural increase

MV ± SD    = Mean value ± Standard deviation

CV             = Coefficient of variation

--               = Not applicable, due to mortality of the parental daphnid

*                 = Adjusted with 1 M NaOH to a pH-value of 7 ± 0.2

¹⁾                =Statistically significant difference (Kruskal-Wallis,One Way Analysis of Variance on Ranks, Dunn’s method, p = 0.05)

Stillborn Juveniles and Aborted Eggs

The number of stillborn juveniles and aborted eggs per surviving parental daphnid of the test and control groups are presented in Table 5.The fraction of dead juveniles (sum of stillborn juveniles and aborted eggs) expressed as a percentage, compared to the total number of produced juveniles (alive, stillborn juveniles and aborted eggs) of the surviving daphnids is presented inTable6.
Related to the total number of produced juveniles (dead + alive) the percentage of dead juveniles corresponded to a maximum of 3 % in the loading levels of 6.25 to 50.0 mg/L and the pH-control. In the loading level of 100 mg/L the percentage of dead juveniles was 8 %. No stillborn juveniles or aborted eggs were observed in the control.

Table5:   Stillborn Juvenilesand Aborted Eggs after 21 Days

Nominal Loading Level of the Test Item [mg/L]	Number of		Total No.		Mean Stillborn Juveniles and Aborted Eggs per Parent alive MV±SD
	Stillborn	Aborted
	Juveniles	Eggs	å	N
100 (pH-control)*	6	14	20	10	15.0 ±5.25
100	36	15	51	9	15.0 ±5.25
50.0	1	10	11	10	15.0 ±5.25
25.0	2	12	14	10	3.10±2.33
12.5	1	29	30	10	1.80±1.62
6.25	0	4	4	10	2.10±2.08
Control	0	0	0	10	0±0

MV ± SD    = Mean value ± Standard deviation

N                = Number of parental daphnids producing juveniles alive

 

Table6:   Relative Number of Dead (Stillborn + Aborted Eggs)to Total Number of Juveniles

Nominal Loading Level of the Test Item [mg/L]	Number of Juveniles			Percentage of Dead Juveniles^#) [%]
	Dead	Alive	Total
100 (pH-control)*	20	972	992	2
100	51	560	611	8
50.0	11	1002	1013	1
25.0	14	978	992	1
12.5	30	862	892	3
6.25	4	952	956	0
Control	0	900	900	0

Dead   = Aborted eggs + stillborn juveniles

Total    = Dead + alive juveniles

^#)          = Related to the total number of juveniles

*          = Adjusted with 1 M NaOH to a pH-value of 7 ± 0.2

 First Appearance of Juveniles

The first day of appearance of juveniles in the replicates producing juveniles of the loading levels 6.25 to 50.0 mg/L, the pH-control and the control was on day 8 or day 9. In the loading level of 100 mg/L four of nine surviving parental daphnids released their first brood after day 10 which is regarded to be biologically significant delayed (Table7).

 

Table7:   First Appearance of Living Juveniles in the Individual Groups

 

Nominal Loading Level of the Test Item [mg/L]	Day of First Appearance of Living Juveniles										First
	in Replicate No.										Appearance
	1	2	3	4	5	6	7	8	9	10	Mean Day
100 (pH-control)*	8	8	9	8	9	8	9	8	8	9	8.4
100	9	9	11^1)	8	--	8	11^1)	8	11^1)	15^1)	10.0
50.0	8	8	8	9	8	8	8	8	8	8	8.1
25.0	8	8	9	8	8	9	8	8	8	8	8.2
12.5	8	8	8	8	8	8	8	8	8	8	8.0
6.25	8	8	8	8	8	9	8	8	8	8	8.1
Control	8	8	8	8	8	8	8	8	8	8	8.0

     --   = Not applicable, due to mortality of the parental daphnid

         ¹⁾   = First brood was biologically significant delayed (after day 10) at 4 of 9 surviving parental daphnids

Adult mortality

No significant adult mortality was observed in the tested loading levels, the pH-control and the control after 21 days. In the loading levels 6.25 to 50.0 mg/L, the pH-control and in the control all parental daphnids survived till the end of the test. In the loading level of 10 mg/L the adult mortality was 10 %, which is not biologically significant. The EL₁₀for adult mortality after 21 days was assessed be 100 mg/L.

Growth (Total Length and Dry Weight) of the Parental Daphnids

At the end of the test the total length and the dry weight of all surviving parental daphnids at each tested loading level, the pH-control and the control were determined (Table9).

The mean values of thebody lengthof the surviving parental daphnids in the treatment groups were determined to be in the range of 5.00 to 5.50 mm per daphnid and 5.33 mm per daphnid at the control group, respectively.
The meandry body weightwas determined to be in the range of 0.71 to 0.95 mg per daphnid in the test groups, 1.14 mg per daphnid in the pH-control, and 0.63 mg per daphnid in the control, respectively.

Presence of Males

No males were observed in either the control or the test groups during the test.

 

 Occurrence of Ephippia (Winter Eggs)

No ephippiawere observedin the control or in the test groups during the test.

 

 Water Quality Parameter

The measured initial and final pH values of the control and the highest tested loading level of 100 mg/L in one replicate per test and control group are presented inTable17.

Significant deviations of > 1.5 units between initial and final pH values were not observed.
The initial pH-values varied from 10.28 to 10.42 in the loading level of 100 mg/L and from 8.02 to 8.26 in the control. The final pH-value varied from 7.19 to 7.33 in the loading level of 100 mg/L and from 7.52 to 7.83 in the control.

The oxygen concentrations of the measured replicates in the test media of the loading level of 100 mg/L and the control were determined to be in the range of 5.19 to 9.44 mg/L (Table18).

The total hardness was determined to be in the range of 134 to 187 mg CaCO₃/L (Table19).

The temperature in the test vessels was within the range 19.1 to 22.0 °C (Table20).

The pH-values of the pH-control were recorded before and after adjustment with 1 M HCl (Table21).

 

 

Acute Immobilization Test of the Reference Item

The percentage immobility for the reference item was determined after 24 h.
The EC₅₀-valueof the reference item with 95 % confidence interval (CI) after 24 h was determined by sigmoidal dose-response regression to be:

                                       EC₅₀          :1.87 mg/L (CI 1.74 - 2.00 mg/L)

The EC₅₀-value of the reference item potassium dichromate after 24 h is within the prescribed concentration range of 1.0 - 2.5 mg/L of quality criteria according toAQS P 9/2 (05/1996) for daphnids clone 5 cultured in Elendt M4 medium. The EC₅₀-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202.

Conclusions:

The Lowest Observed Effect Loading (LOELReproduction) and the No Observed Effect Loading (NOELReproduction) for the reproductive output as the most sensitive effect in this study were deduced directly from the observed decreased reproduction rates in the loading level of 100 mg/L. The EL10-value for the reduction of the reproductive output was calculated by sigmoidal dose-response regression to be 68.2 µg/L. An EL50-value for the reproductive output was not determinable because no effects ≥ 50 % (reduction or increase of the reproductive output) were observed within the tested concentration range.

NOEL Reproduction: 50.0 mg/L
LOEL Reproduction: 100 mg/L
EL10, Reproduction: 68.2 mg/L

Toxic effects were observed on Daphnia magna only in the highest water accommodated fraction* of nominal 100 mg/L. These effects were caused by the high pH-value of > 10. In the pH-control and the nominal loading levels of 6.25 to 50.0 mg/L no effects were observed in comparison to the control. The effect levels given are based on the nominal loading levels.

Executive summary:

The Daphnia magna Reproduction Test (Semi-Static, 21 d) of the test item Flue Dust T(batch No.12-2009) was conducted according to OECD 211 (1998) from2010-08-17 to 2010-09-09with the definitive exposure phase from 2010-08-18 to 2010-09-08 at Dr.U.Noack-Laboratorien, Käthe-Paulus-str.1, 31157,.

Five water accommodated fractions (WAF)* of the test item Flue Dust T were tested over a period of 21 days. Nominal loadings of the test item Flue Dust T for the tested water accommodated fractions* were selected after a preliminary acute immobilization test (static, 48 h) as follows:6.25 - 12.5 - 25.0 - 50.0 - 100 mg/L. Additionally the highest water accommodated fraction* of 100 mg/L was adjusted to a pH-value of 7 ± 0.2 and tested as pH-control.The study was carried out undersemi-static conditions with a three times per week renewal of the test solutions.

For the test Daphnia magna STRAUS (Clone 5) was used. 10 test organisms, individually held, were used per loading level and control. At test start they were 2 to 24 h old. Aim of the Daphnia Reproduction Test over 21 days was to assess effects on the reproduction capacity and other test item-related effects or parameters such as time of production of intrinsic rate of natural increase, first brood, adult mortality, occurrence of aborted eggs and stillborn juveniles and body length of the parental daphnids.

No analytical determination of the test item concentrations in water was carried out. The test item is a mixture of various inorganic compounds. Some of them are nearly insoluble in water. Other compounds dissolve and react according to the well known cement behaviour in water. A single compound which is responsible for any toxic effect cannot be determined. Therefore, a control analysis was not carried out. All effect values given are based on the nominal loading levels of the test itemFlue DustT.

Toxic effects were observed on Daphnia magna only in the highest water accommodated fraction* of nominal 100 mg/L. These effects were caused by the high pH-value of > 10. In the pH-control and the nominal loading levels of 6.25 to 50.0 mg/L no effects were observed in comparison to the control.

 

·  The average number of juveniles per parent alive at the end of the test in the control group was 90.0 after 21 days.The reproductive output was statistically significant reduced in the nominal loading level of 100 mg/L when compared to the control (One Way Analysis of Variance,Dunnett’smethod, p = 0.05).

The EL₁₀-value for the reduction of the reproductive output was calculated by sigmoidal dose-response regression to be 68.2 mg/L. An EL₅₀-value for the reproductive output was not determinable because no effects ≥50 % (reduction of the reproductive output) occurred within the tested concentration range. For details seeTable1, Table3 and Figure1.

  

*)   Water accommodated fraction (WAF): The aqueous medium containing only the fraction of a multi-component

     test item that is dissolved and / or present as a stable dispersion/emulsion under test conditions in the test

     medium, acc. to OECD Series, No. 23.

·  The coefficient of variation of the number of living offspring produced per parent was 9 % in the control. In the loading levels of 6.25 to 50.0 mg/L and the pH-control the calculated coefficients of variation were in the range of 10 to 13 % and comparable to the control. In the loading level of 100 mg/L the coefficient of variation increased to 22 % (Table3).

 

·  The Lowest Observed Effect Loading (LOEL_Reproduction) and the No Observed Effect Loading (NOEL_Reproduction) after 21 days based on the reduction of the reproductive output as the most sensitive effect are summarized inTable1.

 

·  The intrinsic rates of natural increase (IR) of the surviving parent animals accounting for generation time and number of offspring were used for calculation of population growth and maintenance. The mean IR of the surviving daphnids of the treatment groups were compared to the control by Kruskal-Wallis One Way Analysis of Variance on Ranks,Dunn’smethod (p = 0.05). There was a statistical significance in the tested loading level of 100 mg/L when compared to the control. Details are presented in Table1, Table 4 and Figure 2.

 

·  No stillborn juveniles and aborted eggs were produced by the control group. Related to the total number of produced juveniles (dead + alive) the percentage of dead juveniles was in the range of 0 to 3 % in the loading levels of 6.25 to 50.0 mg/L and the pH-control. In the loading level of 100 mg/L the percentage of dead juveniles was 8 % (Table6).

 

·  Four broods were observed within the test period in the loading levels 6.25 to 50.0 mg/L, the pH-control and the control. The first brood was released till day 9 and by all parental daphnids ofthe loading levels 6.25 to 50.0 mg/L, the pH-control and the control.
In the loading level 100 mg/L a biologically significant delay of the first brood after day 10 and less than four broods within the test period was observed at four of nine surviving parental daphnids (Table7).

 

A summary of all test item related effects on reproduction is presented inTable1. The effect levels (NOEL_Reproduction, LOEL_Reproduction and EL_{10 Reproduction}) are given based on the reproductive output as the most sensitive effect.

 

 

Table1:       Test Item Related Effects on Reproduction (NOEL, LOEL,EL_{10, Reproduction})

Effects		Nominal Loading Level of the Test ItemFlue DustT [mg/L]
		Control		6.25		12.5		25.0		50.0		100		100 (pH-control)*
Mean Number of Juveniles per Producing Parent (Reproduction Rate±SD)		90±8		95±10		86±12		98±10		100±10		62±14^1)		97±10
Coefficient of Variation of the Mean Number of Juveniles per Producing Parent [%]		9		10		13		10		10		22		10
Mean Intrinsic Rates of Natural Increase		0.50		0.49		0.51		0.49		0.50		0.40^2^)		0.48
Appearance of First Brood [Mean Day]		8.0		8.1		8.0		8.2		8.1		10.0^3)		8.4
NOELReproduction		50.0 mg/L
LOELReproduction		100 mg/L
EL10, Reproduction		68.2 mg/L
EL50, Reproduction		Not determinable

SD    = Standard deviation

¹⁾      =Statistically significant difference (One Way Analysis of Variance,Dunnett’smethod, p = 0.05)

²⁾      =Statistically significant difference (Kruskal-Wallis,One Way Analysis of Variance on Ranks,Dunn’smethod,
  p = 0.05)

³⁾      = First brood was biologically significant delayed (after day 10) at 4 of 9 surviving parental daphnids

*       =Adjusted with 1 M NaOH to a pH-value of 7 ± 0.2

 

 

A summary of test item related effects concerning the adult mortality and growth (dry weight, body length) is given inTable2.

 

·  Nosignificant adult mortality was observed in the tested loading levels, the pH-control and the control after 21 days. In the loading levels 6.25 to 50.0 mg/L, the pH-control and in the control all parental daphnids survived till the end of the test. In the loading level of 10 mg/L the adult mortality was 10 %, which is not biologically significant. The EL₁₀for adult mortality after 21 dayswas assessedbe 100 mg/L. Details are presented inTable2andTable8.

 

·  The mean values of the body length of the surviving parental daphnids in the treatment groups were determined to be in the range of 5.00 to 5.50 mm per daphnid and 5.33 mm per daphnid at the control group, respectively. The mean dry body weight was determined to be in the range of 0.71 to 0.95 mg per daphnid in the test groups, .1.14 mg per daphnid in the pH-control, and 0.63 mg per daphnid in the control, respectively (Table 2 and Table 9).

 

Table2:   Test Item Related Effects on Mortality and Growthof Parental Daphnids

Effects	Nominal Loading Level of the Test Item Flue Dust T [mg/L]
	Control	6.25	12.5	25.0	50.0	100	100 (pH-control)*
Adult Mortality after 21 Days [%]	0	0	0	0	0	10	0
Mean Dry Weight [mg]	0.63	0.71	0.86	0.83	0.95	0.82	1.14
Mean Body Length [mm]	5.33	5.48	5.43	5.53	5.53	5.00	5.50

*       =Adjusted with 1 M NaOH to a pH-value of 7 ± 0.2

 

·      No males and ephippia (winter eggs) were observed at the control or test groups.

 

·  Water quality parameters as pH-value, dissolved oxygen, water hardness and temperature were determined to be within the acceptable limits. The pH-values of the water accommodated fraction of 100 mg/L were determined to be in the range of pH 10.01 to 10.57. Therefore, a pH-control was tested. Details are presented inTable17toTable21.

 

·  In order to prove the validity of the test system and test conditions at the test facility, an acute immobilization test according to DIN 38412 L 11 is carried out with potassium dichromate as reference item once per month.
The EC₅₀of the reference item at 1.87 mg/L after 24 h is within the prescribed concentration range of 1.0 - 2.5 mg/L of quality criteria according toAQS P 9/2 (05/1996) for daphnids clone 5 cultured in Elendt M4 medium. The EC₅₀-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202. For details see part4.2.

Description of key information

 Toxic effects were observed on Daphnia magna only in the highest water accommodated fraction* of nominal 100 mg/L. These effects were caused by the high pH-value of > 10. In the pH-control and the nominal loading levels of 6.25 to 50.0 mg/L no effects were observed in comparison to the control.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

50 mg/L

Additional information

The Lowest Observed Effect Loading (LOELReproduction) and the No Observed Effect Loading (NOELReproduction) for the reproductive output as the most sensitive effect in this study were deduced directly from the observed decreased reproduction rates in the loading level of 100 mg/L. The EL10-value for the reduction of the reproductive output was calculated by sigmoidal dose-response regression to be 68.2 µg/L. An EL50-value for the reproductive output was not determinable because no effects ≥ 50 % (reduction or increase of the reproductive output) were observed within the tested concentration range.

 

NOEL Reproduction:             50.0 mg/L

LOEL Reproduction:             100 mg/L

EL10, Reproduction:              68.2 mg/L

 

Toxic effects were observed on Daphnia magna only in the highest water accommodated fraction* of nominal 100 mg/L. These effects were caused by the high pH-value of > 10. In the pH-control and the nominal loading levels of 6.25 to 50.0 mg/L no effects were observed in comparison to the control. The effect levels given are based on the nominal loading levels.