The product from the burning of a combination of carbonaceous materials.

Long-term toxicity to fish

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

fish, juvenile growth test

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

Not reported.

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP compliant, non-guideline experimental investigation. Study published in scientific, peer reviewed journal.

Qualifier:

no guideline followed

Principles of method if other than guideline:

Erimyzon sucetta exposed to coal ash were examined for 78 days

GLP compliance:

no

Analytical monitoring:

not specified

Test organisms (species):

other: Erimyzon sucetta

Details on test organisms:

TEST ORGANISM
- Common name: lake chubsuckers
- Source: a historically unpolluted Carolina bay (Bay No. 100), Savannah River Site, SC
- Length at study initiation (length definition, mean, range and SD): 53.56±0.45 mm
- Weight at study initiation (mean and range, SD): 2.76±0.07 g
- Feeding during test: Yes
- Food type: Tetramin
- Amount: 1X, 2X, and 4X i.e. fish assigned to the 1X food level were fed weekly rations of 4.62% of their initial mean body mass (equivalent to 42.5 mg of Tetramin fish food 3 days/week). Fish assigned to the 2X and 4X food levels received weekly rations of 9.24 and 18.48% of their initial mean mass, respectively (equivalent to 85.0 and 170.0 mg of Tetramin fish food 3 days/week). Every 2 weeks, rations of Tetramin in each treatment were increased by 10% of the initial ration in order to accommodate fish growth.

ACCLIMATION
- Acclimation period: 3 weeks
- Type and amount of food: Tetramin fish flakes ad libitum.
- Vehicle: In 72 l tanks containing artificial softwater (US Environmental Protection Agency, 1991)

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 d

Post exposure observation period:

24 h: After 48 h, each post-absorptive fish was placed in an 1100-ml respiratory chamber containing 500 ml of artificial softwater (25 °C) and randomly assigned to a channel on the respirometer. Respiratory chambers were placed in a dark environmental chamber at 25 °C. Oxygen consumption of each fish was determined at 1.5–hour intervals for 24 h. Standard metabolic rate of each fish was estimated as the mean of the lowest 50% of oxygen consumption measurements.

Test temperature:

24.9-25.4°C

pH:

6.74-7.47

Dissolved oxygen:

8.18-8.33 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 60 tanks
- Material, size, headspace, fill volume: glass, 38 L
- Aeration: yes
- Type of flow-through (e.g. peristaltic or proportional diluter): No
- No. of organisms per vessel: 1
- No. of vessels per concentration (replicates): 10 (per level of food)
- No. of vessels per control (replicates): 10 (per level of food)
- Control: sand

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: artificial soft water (US EPA 1991)
- Composition: See table 1.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : mortality 4-5 days per week; growth rate (weight and length) on days 0, 16, 30, 44, 65, and 78; SMR on days 0, 45, and 78; neutral lipids after 78 days.

Reference substance (positive control):

no

Remarks on result:

not measured/tested

Remarks:

Observed parameters included mortality, growth rate (weight and length), SMR and neutral lipids. No LC-, EC-, NOEC or LOEC values were determined.

Details on results:

- Survival rate in 1X was 40% and in 2X and in 4X 90%
- Fish weights (individual and mean values): see figures.
- Other biological observations: erosion on the pectoral and caudal fins, indications of disease

Reported statistics and error estimates:

- dependence (P=0.022) of survival on food level; survival rates of fish were similar among the 2X- and 4X-ash treatments (90% survival), but were low (40%) at the 1X food level.
- dependence of survival on sediment type within the 1X food level (P=0.029)
- patterns of change in fish mass varied significantly among both food and sediment treatments (F2,44=80.30, P<0.001 and F1,44=5.69, P=0.022, respectively).
- significant (P=0.027) correlation of mass with length of fish. Mass was also significantly influenced by food level and the interaction between food level and fish standard length (P=0.014 and 0.013, respectively).
- significant effect of sediment type (P0.001), but no effect of food level (P=0.620), on fin erosion at the end of the study.
- fish SMRs varied significantly through time among sediment type and food level treatment combinations as indicated by the significant time×sediment×food level interaction term in repeated measures ANOVA
- Percent NPL was significantly influenced by food level (F2,44=22.32, P 0.001; Fig. 6)

Table 1 Trace element concentrations in sediments from the site where fish were initially collected and from experimental tanks

	As	Cd	Cu
Collection site	270 ± 0.13	0.40 ± 0.05	16.55 ± 1.24
Control tanks	BDL	BDL	BDL
Ash tanks	58.41 ± 2.99	0.11 ± 0.02	69.12 ± 2.06
Tetramin food	3.93 ± 0.02	0.16 ± 0.01	9.25 ± 0.10

Trace element concentrations for Tetramin fish food are also provided. All trace element concentrations are expressed as mean µg/g dry mass ±SE. For all elements concentrations in reference tanks were below detection limits (BDL). Mean sediment detection limits ( µg/g dry mass) for elements in control and ash tanks (respectively) are as follows, As: 0.408, 0.651; Cd: 0.049, 0.785; Cu: 0.484, 0.722.

Validity criteria fulfilled:

not specified

Conclusions:

Juvenile Erimyzon sucetta exposed to coal ash contaminated sediment showed higher mortality, delay in growth, higher metabolic rate, lower lipid level and fin erosion compared to control fish at lowest nutritional level.

Executive summary:

Sub-chronic effects of coal ash contaminated sediment to juvenile Erimyzon sucetta were studied in a non-GLP compliant non-guideline 78-day static experiment. The exposure time was 48 days, and the post-observation period was 24 days. Aim of the study was to investigate effect of food level on mortality, growth, metabolic rate, and lipid level of the fish. The food levels used were 1X, 2X and 4X, where 1X represented 4.62% of their initial mean body mass. Equal amount of contaminated sediment was used in the test and sand was used as control. In the test tanks, test sediment contained arsenic, cadmium and copper at concentrations between 50-60 µg/g (d.w.). Also, Tetramin fish food had background concentrations below 10 µg/g (d.w.) of these components.The test tanks were aerated and water was filtrated.

Survival rate was lowest (40%) in the group of low nutritional level (1X) compared to other groups, where the rate was 90%. The difference between test sediment and control was significant (P<0.05). Also in group 1X, less nonpolar lipids, lower growth rate and higher metabolic rate was observed. Of these, metabolic rate and fish mass depended on the sediment type. Additionally, fin erosion and indications of disease e.g. red sores, were found in groups 1X and 2X. Change in fish condition was, however, mostly explained by food level (P<0.05). It was discussed, that due to low condition (due to poor nutrition) in group 1X fish, they were not able to tolerate toxic effects.

Reference 2

Endpoint:

fish life cycle toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

No data

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP compliant, non-guideline experimental investigation. Study published in scientific, peer reviewed journal.

Reason / purpose for cross-reference:

reference to same study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Hatchling Cyprinodon variegatus were raised in the presence and absence of sediments contaminated with mixed coal ash over a full life cycle (> 1 yr). Bioaccumulation as well as lethal and sublethal bioenergetic effects (metabolic rate, lipid storage, growth, reproduction) were examined.

GLP compliance:

no

Details on sampling:

On days 75, 112, 166, 184, 238, 266, 303, and 374, one individual fish was arbitrarily selected and removed from each exposure tank for measurement of standard metabolic rate (SMR) by respirometry. Prior to measurement, fish were held individually in freshly mixed water of the proper salinity for 48 h at 25°C, during which time they were unfed. After the measurement fish were blotted dry and weighed before being returned to their original tanks. At the end of the experiment (day 374), all surviving individuals were weighed for wet mass, measured for standard length, classified by sex, and visually examined for fin erosion. Up to three gravid females from each treatment combination were dissected, their eggs were counted, and the egg diameters were measured. One nongravid female and one male from each tank having sufficient survivors were sacrificed and lyophilized prior to analysis for total nonpolar lipid content.

Test organisms (species):

Cyprinodon variegatus

Details on test organisms:

Hatchling C. variegatus (1 day posthatching) were derived from laboratory breeding stocks at the Chesapeake Biological Laboratory (16 ppt) and acclimated to the test salinities (5 or 36 ppt) in a stepwise manner over the next 48 h. Following acclimation, 12 hatchlings were selected arbitrarily for weighing of the initial wet mass. Groups of 12 hatchlings from the acclimation tanks were then randomly placed into aquaria of the corresponding salinity. Fish were fed ad lib a mixture of ground flake and pelletized fish food (1:2 flake:pellet) daily throughout the study.

Test type:

semi-static

Water media type:

saltwater

Total exposure duration:

374 d

Salinity:

The experiments were conducted on two levels of salinity: 5 and 36 ppt.

Nominal and measured concentrations:

Concentrations of the trace elements in contaminated sediment (per dry weight) were: Al: 4179 ppm, As 43.38 ppm, Ba 484.4 ppm, Cd 0.13 ppm and Cu 30.42 ppm. Concentrations of the trace elements in uncontaminated sediments (per dry weight) were: Al: 69 ppm, As 1.76 ppm, Ba 0.02 ppm, Cd 0.036 ppm, Cu 3.26 ppm.

Details on test conditions:

Sediments and water were added to 38-l aquaria arranged randomly in the laboratory. Sediments were added to a depth of 1.5-2 cm in each tank and suspended particles were allowed to settle for a week prior to addition of animals. Tanks were lighted by overhead fluorescent lights on a 12:12 h L:D cycle, and the laboratory was maintained at 23-25°C. Salinity, temperature and dissolved oxygen conditions were monitored in each tank on a weekly basis. Fish were fed ad lib a mixture of ground flake and pelletized fish food (1:2 flake:pellet) daily throughout the study. At approximately 50-75 day intervals, tanks were drained and cleaned, and the sediments and water were replaced. Fish removed from the tanks during cleaning were blotted dry and weighed before being returned to the tanks.

Key result

Duration:

374 d

Dose descriptor:

other: See details on results

Effect conc.:

other: Al: 4179 ppm, As 43.38 ppm, Ba 484.4 ppm, Cd 0.13 ppm and Cu 30.42 ppm

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Remarks:

sediment

Basis for effect:

other: growth, standard metabolic rate, storage lipid content, size of eggs

Remarks on result:

other: Effect concentrations: Al: 4179 ppm, As 43.38 ppm, Ba 484.4 ppm, Cd 0.13 ppm and Cu 30.42 ppm

Details on results:

Growth (change in wet mass) throughout the duration of the study was significantly higher for control animals than those exposed to contaminants. Growth effects became apparent in the latter half of the experiment (from about day 200 onward). Average final wet masses of females and males exposed to contaminated sediment were, respectively, about 30% and 37% lower than wet masses of controls. Standard lengths were 10–11% lower for both sexes following contaminant exposure compared to control fish. While differences in final wet mass and standard length of each sex between treatments were not significant, the differences certainly may be biologically significant. Condition factor, an integrated measurement of mass and length, was significantly reduced in the contaminated treatment compared to the control treatment for males, but not for females. Lipid storage by females was lower in contaminated than in control treatments, but male lipid content did not differ between treatments. SMR followed the predicted pattern, being initially high in the rapidly growing and developing juveniles and decreasing with time. However, throughout the study SMR did not differ significantly between contaminant-exposed and control animals. At the end of the experiment, there was no statistically significant difference between treatments in the proportion of females that were gravid with ripe eggs nor in the number of ripe eggs per gravid female. However, eggs removed from females in the control treatment were significantly larger (by about 12%) than those removed from females raised in the presence of contaminated sediments. No individuals displayed fin erosion. Salinity had no effect on responses measured.

Reported statistics and error estimates:

- Growth (change in wet mass) throughout the duration of the study (from ANOVA of repeated observations) was significantly higher for control animals than those exposed to contaminants ( F1,68 = 5:20; P=0.038).
- Thus the slope of the growth trajectory was reduced in
contaminated sediment treatments compared to controls
(95% confidence intervals for slopes of growth trajectories
for contaminant and control treatments, respectively:
0.002970.0005 and 0.004970.0011).
- Differences in final wet mass and standard length of each sex between treatments were not significant at the a priori Type I error rate of a = 0.05
(P =0.0562 - 0.104)
- SMR did not differ significantly between contaminant-exposed and control animals (F1,99 = 1.67; P = 0.213)

Table 1 Responses of C. variegatus following an exposure to trace element-contamination or control sediments, from hatchling through adulthood (374 days)

Survival (%)

Wet mass, females (g)

Wet mass, males (g)

Standard length, females (cm)

Standard length, males (cm)

Condition factor, females (g/cm^3)

Condition factor, males (g/cm^3)

Nonpolar lipid content, females (%)

Nonpolar lipid content, males (%)

Gravid females (%)

Number of ripe eggs

Diameter of ripe eggs (mm)

Contiminated sediment

56±9

0.65±0.07

1.12±0.15

2.82±0.10

3.36±0.14

0.029±0.001

0.029±0.001

13.3±1.8

11.5±3.1

22.2±11.1

4.0±2.9

1.22±0.07

Control sediment

51±8

0.94±0.12

1.76±0.27

3.16±0.15

3.72±0.15

0.029±0.001

0.033±0.002

16.5±1.4

16.4±3.9

50.0±14.5

3.7±0.9

1.39±0.11

F^1,12= 0.17

F^1,12= 4.37

F^1,12= 4.47

F^1,12= 3.61

F^1,12= 3.09

F^1,12= 0.34

F^1,12= 7.26

F^1,14= 6.34

F^1,9= 2.11

F^1,11= 2.58

F^1,10= 0.01

F^1,7= 5.70

P=0.691

P=0.059

P=0.056

P=0.082

P=0.104

P=0.571

P=0.019

P=0.033

P=0.180

P=0.137

P=0.914

P=0.048

Validity criteria fulfilled:

not specified

Conclusions:

Over an exposure period lasting over the entire life cycle of the C. variegatus, reductions in growth, condition factor, lipid storage content, and size of eggs produced resulted from trace element contamination of sediments. However, the biological effects were slow to emerge, being observed in individuals in the late juvenile and adult life stages, rather than in the presumably more sensitive early life stages.

Executive summary:

Long-term toxicity of coal ash contaminated sediment to hatchling Cyprinodon variegatus was measured in a non-GLP, non-guideline experimental study. C. variegatus were raised in the presence or absence of sediments contaminated with mixed trace elements to examine lethal and sublethal bioenergetic effects (metabolic rate, lipid storage, growth, reproduction) over a full life cycle (>1 year). Contaminated sediments were derived from a site receiving coal combustion residues and contained elevated concentrations of numerous trace elements including Al, As, Ba, Cd, and Cu.

There were no differences in fish survival for contaminated sediment treatments and uncontaminated sediment treatments, nor were there differences in metabolic expenditures. However, growth, male condition factor, and storage lipid content in females were reduced due to contaminant exposure. No significant effects on fecundity or the proportion of females that were gravid at the end of the study were observed, yet females raised under control conditions produced 12% larger eggs than did females raised on contaminated sediments. During the presumably most-sensitive early life stages, individuals were not noticeably affected by contaminants, but the effects of exposure became apparent later in life.

Description of key information

Long-term toxicity to fish was estimated based on two publications. The studies were non-GLP compliant non-guideline experimental investigations. In the first study, juvenile Erimyzon sucetta was exposed to coal ash contaminated sediment in a 78-h subchronic study with different nutritional levels also included in the test. The toxic effects i.e. mortality, growth delay and higher metabolic rate were found to depend significantly on the level of nutrition (p<0.05). Additionally, fin erosion was reported. With normal food level, effects except delay in growth were not observed compared to control. In the other study, toxicity of coal ash contaminated sediment to hatchling Cyprinodon variegatus was estimated in a long-term test covering a full life cycle (>1 year). There were no differences in fish survival for contaminated sediment treatments and uncontaminated sediment treatments, nor were there differences in metabolic expenditures. However, growth, male condition factor, and storage lipid content in females were reduced due to contaminant exposure. No significant effects on fecundity or the proportion of females that were gravid at the end of the study were observed, yet females raised under control conditions produced 12% larger eggs than did females raised on contaminated sediments. During the presumably most-sensitive early life stages, individuals were not noticeably affected by contaminants, but the effects of exposure became apparent later in life.

Additional information

Data on EC10/LC10 or NOEC is lacking. As a conclusion on the basis of the publications, ash contamination of water ecosystem can cause delay in growth, but does not have negative influences on breeding and vitality of the offspring. Effects on the breeding of the next generation and later generations cannot be concluded.