Ashes (residues), cenospheres

Toxicity to reproduction

Administrative data

Endpoint:

fertility, other

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

10 Dec 1950 - 14 Apr 1954

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well-documented publication which meets basic scientific principles.

Cross-reference

Reason / purpose:

reference to same study

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Unnamed

Year:

1955

Reference 2

Reference Type:

publication

Title:

Fütterungsversuch mit Steinkohlenflugasche bei Milchkühen.

Author:

Goetze, R., und Herrmann, M.

Year:

1956

Bibliographic source:

Schriftenreihe des Vereins für Wasser-, Boden- und Lufthygiene Berlin-Dahlem 11, 81-219 (1956), Gustav Fischer Verlag, Stuttgart.

Reference 3

Reference Type:

publication

Title:

Die Nutzbarkeit der Steinkohlenflugasche zur Mineralstoffversorgung der Haustiere.

Author:

Goetze, R.

Year:

1956

Bibliographic source:

Schriftenreihe des Vereins für Wasser-, Boden- und Lufthygiene Berlin-Dahlem 11, 221-224 (1956), Gustav Fischer Verlag, Stuttgart.

Materials and methods

Principles of method if other than guideline:

Fertility study in grazing cows

GLP compliance:

not specified

Limit test:

no

Test material

Reference

Name:

Unnamed

Type:

Constituent

Details on test material:

- Name of test material (as cited in study report): Fly ash from (black) coal combustion, Scholven, Germany
- Other: Further details are given in the tables below

Test animals

Species:

other: cattle

Strain:

other: Schwarzbunte Niederungsrasse

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Oldenburg, Germany (a geographical region which was assumed to be free of fly ash from coal combustion)
- Age at study initiation: 2.75 - 6.75 years
- Weight at study initiation: 535 - 710 kg
- Housing: Cattle pen; the animals were allowed to graze.
- Diet (e.g. ad libitum): fodder beed, hay and straw (ad libitum) supplemented with mash.
At their arrival the animals were carefully clinically examined. All animals were in good nutritional state and showed no symptoms of illness.
Some animals were seropositive for tuberculosis.

ENVIRONMENTAL CONDITIONS
All animals were kept on farm without contact to other cattle.
The cattle pen was always kept clean, dry and ventilated with fresh air.

IN-LIFE DATES:
Exposure period: From: 10 Dec 1950 To: 15 Dec 1953
Post exposure observation period: From: 16 Dec 1953 To: 14 Apr 1954

Administration / exposure

Route of administration:

oral: feed

Vehicle:

unchanged (no vehicle)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
The test substance was mixed with the daily food.
Animals were fed twice a day in individual boxes to assure the intake of the individual portions.

DIET PREPARATION
In the high dose group, the fly ash was mixed with soaked fodder beed and mash, to improve the taste and to ascertain the daily intake of the high amounts of fly ash.

Details on mating procedure:

All animals were naturally mated.
At the beginning of the experiment, 8 out of 9 animals were already pregnant.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

Exposure period: 3 years
Premating exposure period (females): no data

Frequency of treatment:

twice a day

Details on study schedule:

not applicable

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
300 g / animal / day
Basis:
nominal conc.
fed to the animals of group 1 and group 2

Doses / concentrations 2

Remarks:

Doses / Concentrations:
1500 - 1800 g / animal / day
Basis:
nominal conc.
fed to the animals of group 3

Doses / concentrations 3

Remarks:

Doses / Concentrations:
additionally approx. 300 g / animal / day
Basis:
nominal conc.
resulting from calculated amounts of coal fly ash precipitation to the grazing land in the experimental geographic area and the daily intake of the grazing test animals.

No. of animals per sex per dose:

9 cows were split into 3 groups (3 animals/group):
Group 1 (animal # 1-3): served as concurrent no treatment control animals in the first two years and received 300 g fly ash/day/animal in the 3rd year of the experiment
Group 2 (animal # 4-6): received 300 g fly ash/day/animal only in the 1st and 2nd year of the experiment, in the 3rd year concurrent no treatment.
Group 3 (animal # 7-9): received 1500 - 1800 g fly ash/day/animal during all 3 years of the experiment

Control animals:

other: yes, concurrent no treatment (for the first two years of the exposure period)

Details on study design:

To exclude possible individual differences in the sensitivity for coal fly ash, a feeding strategy conversion for animal groups 1 and 2 was performed: Animals of group 1 received fly ash only in the third year, whereas the animals of group 2 received the fly ash only in the first two years of the 3-year duration.

Positive control:

not performed

Examinations

Parental animals: Observations and examinations:

REGULAR CLINICAL OBSERVATIONS: Yes
- Time schedule:
First year: every 10 days
Second year: every 14 days
Third year: every 4 weeks

DETAILED CLINICAL OBSERVATIONS: Yes, including parasitological, bacteriological and serological analyses
- Time schedule:
1) at the arrival
2) at the end of the second year of exposure
3) at the end of the exposure period of 3 years
4) at the end of the post-exposure period

BODY WEIGHT: Yes
- Time schedule for examinations:
First year: every 10 days
Second year: every 14 days
Third year: every 4 weeks

MILK PRODUCTION:
Every day the milk production was reported by weighing.

OTHER:
Milk, blood, urine and faeces samples for chemical analyses were taken on December 10th, 1952.
Trace element analysis of liver and bone samples were performed at the end of the experiment.

Sperm parameters (parental animals):

not analysed

Litter observations:

The parturition of all calves was described as undisturbed, but the calves were not further mentioned in the report.

Postmortem examinations (parental animals):

SACRIFICE
- Maternal animals: 2 animals of each group were sacrified at the end of the exposure period, the remaining animals were euthanised at the end of the post exposure observation period.

GROSS NECROPSY
All animals were macroscopically examined.

HISTOPATHOLOGY / ORGAN WEIGHTS
Samples from : Liver, kidneys, pyloric region of the stomach, mesenterial lymph nodes, small and large intestine

OTHER:
Liver samples were chemically analysed for their trace element contents.

Postmortem examinations (offspring):

not examined

Statistics:

not applicable (n=3)

Reproductive indices:

not examined

Offspring viability indices:

not examined

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Other effects:

no effects observed

Details on results (P0)

see tables below

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

1 850 - 2 390 mg/kg bw/day (nominal)

Sex:

female

Basis for effect level:

other: overall effects clinical signs; mortality; body weight; gross pathology; histopathology; pregnancy rate dose level calculated on the basis of the reported body weight range (627-809 kg)

Results: F1 generation

General toxicity (F1)

Mortality / viability:

no mortality observed

Overall reproductive toxicity

Reproductive effects observed:

not specified

Any other information on results incl. tables

Table 1: Details on the experimental settings

Exposure period: December 10, 1950 - December 15, 1953;

Post exposure recovery period: December 16, 1953 - April 14, 1954

Group	Animal numbers	Treatment during the 1^stand 2^ndyear (until December 10, 1952)	Treatment during the 3^rdyear
A	1-3	concurrent no treatment	300 g fly ash / day / animal
B	4-6	300 g fly ash / day / animal	concurrent no treatment
C	7-9	1500 - 1800 g fly ash /day / animal	1500 - 1800 g fly ash /day / animal

Applicant's conclusion:

Only in the first two years of the experiment, group A served as a control group. Therefore, as far as possible only results after two years of experimental duration were evaluated.

 

Table 2: Individual observations

Ani-mal	reco-very after expo-sure	Age, body weight and preg-nancy status at the experi-mental start	Clinical signs	Preg- nancy rate^a	Sexual behaviour	Body weight (in kg at similar stages of preg- nancy, measured in 1950, 1951, 1952 and 1953)	Milk produc-tion  (kg / d; mean values of the years 1951 and 1952)
1	no	5 years, 657 kg, 5^thmonth	No clinical signs of toxico- logical relevance were observed.	4/6	No disturbances of conception and parturition.	657, 697, 720, 744	10.9
2	yes	2.75 years, 535 kg, 9^thmonth	No clinical signs of toxico- logical relevance were observed.	4/4	No disturbances of conception and parturition up to the 3^rdbirth in 1953. During the 4^thpregnancy an infection with Brucella abortus occured, followed by a preterm delivery in the 8^thmonth of pregnancy.	535, n.d., 590, 682	10.7
3^b	no	5 years, 585 kg, not pregnant	No clinical signs of toxico- logical relevance were observed.	3/3	No disturbances of conception and parturition up to the 2^ndbirth in April, 1953. In November 1953 an abortion occured after a Brucella infection.	n.d., n.d., 554, 709	11.6
4	yes	4.75 years, 625 kg, 5^thmonth	Observed clinical signs (uterus catarrh, bronchial catarrh) couldn´t be associated with fly ash administration, as the milk production was highest during fly ash administration	3/6	Mastitis due to a Streptococcus infection occured early after experimental start. Infection with Brucella abortus during post exposure period.	625, n.d., 695, 691	11.3
5	no	4.5 years, 645 kg, 9^thmonth	No clinical signs of toxico- logical relevance were observed.	4/6	No disturbances of conception and parturition up to the 3^rdbirth in 1953. During the 4^thpregnancy an infection with Brucella abortus occured, followed by an abortion in the 8^thmonth of pregnancy.	645, 636, 686, n.d.	13.7
6	no	5 years, 710 kg, 9^thmonth	No clinical signs of toxico- logical relevance were observed. The occuring mastitis could not be associated with fly ash administration.	4/4	No disturbances of conception and parturition up to the 3^rdbirth. During the 4^thpregnancy the animal was euthanised after an accident with fracturing of the lumbar vertebrae.	710, 727, n.d., 812	12.9
7	no	5.5 years, 627 kg, 7^thmonth	No clinical signs of toxico- logical relevance were observed.	4/4	Abortion in February 1951 without evidence of bacterial infection. No disturbances of conception and parturition were observed for the following 3 pregnancies.	627, 683, 685, n.d.	11.9
8	no	5.75 years, 670 kg, 7^thmonth	No clinical signs of toxico- logical relevance were observed.	4/7	No disturbances of conception and parturition up to the 3^rdbirth. During the 4^thpregnancy the animal was euthanised.	670, 719, 732, n.d.	10.3
9	yes	6.75 years, 682 kg, 9^thmonth	No clinical signs of toxico- logical relevance were observed.	4/4	No disturbances of conception and parturition during all 4 pregnancies and deliveries.	682, 765, 784, 809	12.3

n.d. = not determined

^anumber of pregnancies / number of natural matings

^bAnimal #3 was incorporated to the experiment in March 1952 as a replacement for another animal, which was euthanised due to diseases of the lung and intestine. This was considered as not relevant, as this animal belonged to the concurrent no-treatment group.

 

Applicant's conclusion:

Various confounding factors like several infectious diseases like Brucella abortus, Tuber-culosis, liver Trematodes and others (viral infections were not mentioned at all) and complications following accidental injuries (following natural matings, for instance) exacerbate the interpretation of the individual observations. However, no dose-dependent changes of the sexual behaviour, pregnancy rate or development of conceptus and parturition were noted. The general condition of the animals seemed to be not disturbed by the test substance administration, as the milk production was even slightly higher in both treated groups compared to the control group.

Body weights of the high dose group was ranged from 627 to 809 kg. Accordingly, the applied dose was ca. 1850 -2390 mg fly ash / kg body weight / day in the high dose group.

 

Table 3:Individual macroscopic and microscopic pathological findings

Ani- mal / Treat- ment	Necropsy findings	Histological findings^ain kidney, liver, stomach, lymph nodes and intestine samples
1	infectious diseases: Liver-Trematodes, Tuberculosis; changes in claws, which were considered as normal in this species under the given circumstances; no irritation of gastro-oesophageal mucosae, no changes in bones and joints^b, which could be a consequence of a fly ash intoxication;	no histopathologic changes observed
2	infectious diseases: Liver-Trematodes, Tuberculosis, Brucella abortus; changes in claws, which were considered as normal in this species under the given circumstances; no irritation of gastro-oesophageal mucosae, no changes in bones and joints	liver: high glycogen deposition and fatty changes; kidney: glomerulonephritis, brownish pigmentation in tubular epithelial cells, mild nephrosis; lymph nodes: pigmentation
3	infectious diseases: Liver Trematodes, Brucella abortus; no irritation of gastro-oesophageal mucosae; changes in uterus and vagina were considered to be a consequence of Brucella infection; no changes in joints;	no histopathologic changes observed
4	infectious diseases: Tuberculosis, Streptococcus, Brucella abortus; no irritation of gastro-oesophageal mucosae; no changes in joints;	kidney: glomerulonephritis, pigmentation of tubular epithelial cells; liver: fatty changes, glycogen deposits in centrolobular cells.
5	infectious diseases: Liver Trematodes, Tuberculosis, Brucella abortus; petechial changes of the gall bladder mucosa, which were considered as not toxicologically relevant; no irritation of gastro-oesophageal mucosae; no changes in joints;	no histopathologic changes observed
6	infectious diseases: Tuberculosis; fracture of the lumbar vertebra; a necropsy of this animal was not considered, as the animal suffered for 6 weeks from the downer cow syndrome due to the lumbar vertebra fracture.	kidneys: lymphocytic interstitial infiltration; liver: mild perivascular lymphocytic infiltration;
7	infectious diseases: Liver Trematodes; mild erythema^cin the pyloric mucosa of the abomasum, the amounts of sand in the gastro-oesophageal mucosa was considered as normal, no defects, bleeding or necrosis in the mucosa of the gastro-oesophageal tract, no other findings of toxicological relevance; no changes in bones and joints;	no histopathologic changes observed with exception for a focal fatty change (fine drops in only one place in the liver)
8	infectious diseases: Liver Trematodes; the amounts of sand in the gastro-oesophageal mucosa was considered as normal; no irritation of gastro-oesophageal mucosae; no changes in bones and joints;	no histopathologic changes observed
9	infectious diseases: Liver Trematodes; the amounts of sand in the gastro-oesophageal mucosa was considered as normal; mild erythema in the abomasal mucosa, no defects, bleeding or necrosis in the mucosa of the gastro-oesophageal tract, no changes in bones and joints;	kidney: chronic glomerulonephritis, focal nephritis chronica interstitialis, protein deposition in tubular epithelial cells; liver: hydropic degeneration with cell lysis, mild fatty change and glycogen deposits in the centrolobular region; lymph nodes: pigmentation

^aThe histologic findings detected in animals #2, #4 and #9 (groups A, B and C) were considered to be without toxicological relevance, as these finding were without a dose-response relation.

^bThe joints which were previously described to be affected in animals chronically intoxicated with fly ash

^cThe mild erythemas in some gastrooesophageal tracts were considered to be of no toxicological relevance, as this finding was also observed in 25 out of 30 animals analysed at the butchery of Hannover, Germany.

 

Applicant's conclusion:

The pathological findings observed after 3 years of experimental duration are of low toxicological relevance, as too many known and unknown confounding factors like infectious diseases were present and due to the fact, that the treatment regime of animal groups A and B were exchanged after 2 years. However, no major pathological defects of the gastro-intestinal mucosa or other target organs like the joints occurred. Therefore it can be summarized that oral administration of the fly ash up to 1850 mg/kg/day to female cattle for three years did not result in major pathological changes.

 

Table4: Analysis of milk samples collected on December 10, 1952:

(Average values in % determined from two different laboratories in parallel)

Animal	Treatment up to the sampling date	Dry residue	Ash	CaO	MgO	P2O5
1	concurrent no treatment	12.9	0.74	0.172	0.0144	0.212
2	concurrent no treatment	15.8	0.89	0.267	0.0222	0.276
3	concurrent no treatment	10.3	0.71	0.136	0.0155	0.185
4	300 g fly ash per day	11.2	0.78	0.173	0.0170	0.235
5	300 g fly ash per day	14.7	0.82	0.197	0.0194	0.299
6	300 g fly ash per day	14.1	0.71	0.152	0.0204	0.159
7	1500 - 1800 g fly ash per day	15.3	0.69	0.161	0.0161	0.220
8	1500 - 1800 g fly ash per day	14.1	0.73	0.155	0.0144	0.213
9	1500 - 1800 g fly ash per day	10.3	0.73	0.151	0.0153	0.224

Applicant's conclusion:

Two years of oral administration of two different doses of fly ash did not influence the composition of the milk regarding the analysed parameters.

 

Table5: Analysis of blood samples collected on December 10, 1952:

(Average values in % determined from two different laboratories in parallel)

Animal	Treatment up to the sampling date	Dry residue	Ash	CaO	MgO	P2O5	Fe2O3
1	concurrent no treatment	19.2	1.37	0.0127	0.0058	0.0374	0.0534
2	concurrent no treatment	19.9	1.03	0.0110	0.0060	0.0380	0.0544
3	concurrent no treatment	18.7	1.15	0.0116	0.0061	0.0371	0.0435
4	300 g fly ash per day	19.5	1.22	0.0133	0.0062	0.0396	0.0502
5	300 g fly ash per day	19.2	1.36	0.0094	0.0077	0.0333	0.0572
6	300 g fly ash per day	19.6	1.13	0.0101	0.0068	0.0350	0.0430
7	1500 - 1800 g fly ash per day	20.3	1.16	0.0105	0.0071	0.0359	0.0480
8	1500 - 1800 g fly ash per day	18.3	0.98	0.0112	0.0062	0.0362	0.0489
9	1500 - 1800 g fly ash per day	19.3	1.12	0.0116	0.0077	0.0345	0.0547

Applicant's conclusion:

Two years of oral administration of two different doses of fly ash did not influence the composition of the blood regarding the analysed parameters.

 

Table 6: Analysis of urine samples collected on December 10, 1952

(Average values in % determined from two different laboratories in parallel)

Animal	Treatment up to the sampling date	pH	Glucose	Protein	Dry residue	CaO	MgO^a	P2O5
1	concurrent no treatment	8.14	neg.	neg.	6.84	0.0055	0.0170	0.0027
2	concurrent no treatment	8.20	neg.	neg.	5.18	0.0018	0.0221	0.0024
3	concurrent no treatment	8.18	neg.	neg.	5.41	0.0011	0.0261	0.0016
4	300 g fly ash per day	8.36	neg.	neg.	5.33	0.0066	0.0090	0.0013
5	300 g fly ash per day	8.26	neg.	neg.	4.53	0.00097	0.0080	0.0014
6	300 g fly ash per day	8.34	neg.	neg.	5.11	0.0043	0.0302	0.0031
7	1500 - 1800 g fly ash per day	8.08	neg.	neg.	3.85	0.0084	0.0100	0.0013
8	1500 - 1800 g fly ash per day	8.40	neg.	neg.	6.33	0.0041	0.0357	0.0022
9	1500 - 1800 g fly ash per day	8.38	neg.	neg.	5.80	0.0233	0.0411	0.0025

^aMgO was only analysed from one laboratorium

Applicant's conclusion:

Two years of oral administration of two different doses of fly ash did not influence the composition of the urine regarding the analysed parameters.

 

Table8: Analysis of faeces sampled on December 10, 1952

(Values in % of fresh faeces)

Animal	Treatment up to the sampling date	Ash	SiO2	Al2O3	Fe2O3	MgO	CaO	SO3	P2O5
1	concurrent no treatment	2.77	1.51	0.119	0.066	0.118	0.270	0.057	0.183
2	concurrent notreatment	2.23	1.11	0.086	0.054	0.150	0.250	0.044	0.164
3	concurrent no treatment	2.28	1.13	0.091	0.062	0.111	0.259	0.052	0.167
4	300 g fly ash per day	2.68	1.32	0.296	0.142	0.131	0.255	0.054	0.190
5	300 g fly ash per day	2.36	1.17	0.240	0.148	0.111	0.240	0.054	0.170
6	300 g fly ash per day	2.98	1.55	0.431	0.201	0.131	0.350	0.077	0.270
7	1500 - 1800 g fly ash per day	4.81	2.49	0.716	0.615	0.286	0.372	0.107	0.182
8	1500 - 1800 g fly ash per day	6.21	2.84	0.827	0.643	0.352	0.477	0.135	0.246
9	1500 - 1800 g fly ash per day	4.76	2.31	0.644	0.506	0.288	0.492	0.105	0.208

^aMgO was only analysed from one laboratorium

Applicant's conclusion:

Two years of oral administration of two different doses of fly ash influenced the ash and mineral contents of the faeces in a dose-dependent manner.

 

Analysis of trace elements in liver and bone samples:

Liver and bone samples of all animals were analysed for their contents of Arsenic, Manganese, Lead, Iron, Copper, Zinc and Cobalt. Bones were additionally analysed for their CaO and P₂O₅content.

In summary, there was no significant influence of the fly ash administration on the trace element contents in liver and bone samples.

Applicant's summary and conclusion

Conclusions:

A NOAEL for female cattle of ca. 1850-2390 mg/kg bw/day could be identified. Male reproduction parameters were not examined. Litters were not examined.