Registration Dossier

Diss Factsheets

Use of this information is subject to copyright laws and may require the permission of the owner of the information, as described in the ECHA Legal Notice.

REACH

Manganese di(acetate)

EC number: 211-334-3 | CAS number: 638-38-0

Life Cycle description
Uses advised against

Toxicological information

Repeated dose toxicity: oral

Currently viewing:

Administrative data

Endpoint:: chronic toxicity: oral
Remarks:: combined repeated dose and carcinogenicity
Type of information:: migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:: key study
Study period:: from 24 September 1984 to 10 October 1986
Reliability:: 2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:: other: Read-across substance, performed similar to corresponding guidelines, well documented report including raw data.

Data source

Reference

Reference Type:: study report
Title:: Unnamed
Year:: 1993

Materials and methods

Test guideline

Qualifier:: equivalent or similar to guideline
Guideline:: OECD Guideline 453 (Combined Chronic Toxicity / Carcinogenicity Studies)

GLP compliance:: not specified

Test material

Test material information

Details on test material:: - Name of test material (as cited in study report): Manganese (II) sulfate monohydrate
- Molecular formula (if other than submission substance): MnSO4*H2O
- Substance type: pure substance
- Physical state: solid
- Analytical purity: 97.7% ± 0.4%, determined by the analytical chemistry laboratory, Midwest Research Institute (Kansas City, MO)
- Impurities (identity and concentrations): sodium (640 ppm), potassium (120 ppm), and silicon (160 ppm)
- Lot/batch No.: 003261
- Other: white, slightly efflorescent crystalline compound

Test animals

Species:: other: rats and mice
Strain:: other: F344/N rats and B6C3F1 mice
Sex:: male/female
Details on test animals or test system and environmental conditions:: TEST ANIMALS
- Source:
Frederick Cancer Research Facility (Frederick, MD, USA)
- Age at study initiation:
41 d (2a study, mice)
41 d (2a study, rats)
- Weight at study initiation:
rats, male, 2a: 123 - 126 g
rats, female, 2a: 100 - 104 g
mice, male, 2a: 20.6 - 21.1 g
mice, female, 2a: 17.0 - 17.1 g
- Fasting period before study:
- Housing:
Cages: Polycarbonate (Lab Products, Inc., Garfield, NJ, USA), changed twice weekly
Bedding: BetaChips, hardwood chips (Northeastern Products, Inc., Warrensburg, NY), changed twice weekly or more frequently when needed.
Rats and mice were housed five per cage. Feed and water were available ad libitum. Feed consumption was measured for 7 days, once a month. Cages were rotated every 2 weeks; racks were rotated every 2 weeks.
- Diet (e.g. ad libitum): NIH-07 (60 g manganese oxide per 2,000 lbs feed) open formula meal rat and mouse diet (Zeigler Brothers, Inc., Gardners, PA, USA), available ad libitum
- Water (e.g. ad libitum): Automatic watering system (Edstrom Industries, Waterford, WI), available ad libitum
- Acclimation period:
13 d (2a study, mice)
12 d (2a study, rats)
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.6° - 23.9° C
- Humidity (%): 35 - 65%
- Air changes (per hr): min. 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:

From: No data.

To:

Rats:
9-Month interim: 25-26 June 1985
15-Month interim: 4 December 1985
Terminal: 22-25 September 1986

Mice:
9-Month interim: 10-11 July 1985
15-Month interim: 2-3 January 1986
Terminal: 6-10 October 1986

Administration / exposure

Route of administration:: oral: feed
Vehicle:: unchanged (no vehicle)
Details on oral exposure:: PREPARATION OF DOSING SOLUTIONS:

DIET PREPARATION
- Rate of preparation of diet (frequency): Maximum Storage Time: 21 days from date of preparation, preparation weekly
- Mixing appropriate amounts with (Type of food): NIH-07 (60 g manganese oxide per 2,000 lbs feed) open formula meal rat and mouse diet (Zeigler Brothers, Inc., Gardners, PA, USA)
- Storage temperature of food: 25°C
Analytical verification of doses or concentrations:: yes
Details on analytical verification of doses or concentrations:: Periodic monitoring of the chemical by the study laboratory using chelometric titration and elemental analyses (Galbraith Laboratories, Inc., Knoxville, TN) indicated no degradation of the bulk chemical during the studies
Duration of treatment / exposure:: 103 weeks
Frequency of treatment:: Test substance is applied via feed, available ad libitum

Doses / concentrations

Remarks:: Doses / Concentrations:
0, 1.500, 5.000, or 15.000 ppm in feed, available ad libitum
Basis:
nominal in diet

No. of animals per sex per dose:: 70 males and 70 females
Control animals:: yes, plain diet
Details on study design:: - Dose selection rationale:
Based on decreases in body weight gain and the lower absolute and relative liver weights in the 25,000 ppm groups in the 13-week study, doses of 0, 1,500, 5,000, and 15,000 ppm were selected for the 2-year study in rats.
The doses selected for the 2-year study in mice were 0, 1,500, 5,000, and 15,000 ppm. These doses were based on the significantly lower mean body weight gains of all exposed males and 50,000 ppm females and the significantly lower absolute and relative liver weights of 50,000 ppm males in the 13-week study

Examinations

Observations and examinations performed and frequency:: CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: initially, weekly during first 13 weeks of study, monthly thereafter

BODY WEIGHT: Yes
- Time schedule for examinations: initially, weekly during first 13 weeks of study, monthly thereafter

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No. Feed consumption measured for a 7-day period once every 4 weeks.
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No.
Ad libitum.

FOOD EFFICIENCY: No

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No.

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: at the 9- and 15-month
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: All
- Parameters checked in table G3, G4, G7, G8 were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: at the 9- and 15-month
- Animals fasted: No data
- How many animals: All

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No
Sacrifice and pathology:: GROSS PATHOLOGY: all organs and tissues were examined for gross lesions
HISTOPATHOLOGY: Yes: and all major tissues were fixed and preserved in 10% neutral buffered formalin, processed and trimmed, embedded in paraffin, sectioned to a thickness of 5 to 6 µm, and stained with hematoxylin and eosin for microscopic examination. Complete histopathologic examinations were performed on 0 and 15,000 ppm animals at the 9- and 15-month interim evaluations and gross lesions were examined for the 1,500 and 5,000 ppm groups. Complete histopathologic examinations were performed on all animals surviving until the end of the studies and on those that died or were killed moribund during the studies.

Results and discussion

Results of examinations

Clinical signs:: no effects observed
Description (incidence and severity):: Rats: Survival of 1972 mg/kg bw male rats in the 2-year study was significantly lower than that of the control group. The deaths of males in the control and exposure groups were attributed to a variety of spontaneous neoplastic and nonneoplastic lesions; however, the greater number of deaths in the 1972 mg/kg bw group resulted from increased incidences of advanced renal disease related to ingestion of manganese (II) sulfate monohydrate. Mice: No significant effect.
Mortality:: no mortality observed
Description (incidence):: Rats: Survival of 1972 mg/kg bw male rats in the 2-year study was significantly lower than that of the control group. The deaths of males in the control and exposure groups were attributed to a variety of spontaneous neoplastic and nonneoplastic lesions; however, the greater number of deaths in the 1972 mg/kg bw group resulted from increased incidences of advanced renal disease related to ingestion of manganese (II) sulfate monohydrate. Mice: No significant effect.
Body weight and weight changes:: no effects observed
Description (incidence and severity):: Rats and mice: no significant effect compared to control.
Food consumption and compound intake (if feeding study):: no effects observed
Food efficiency:: not specified
Water consumption and compound intake (if drinking water study):: not specified
Ophthalmological findings:: not examined
Haematological findings:: effects observed, treatment-related
Description (incidence and severity):: Rats: Slight differences between exposed and control groups were not considered related to the ingestion of manganese (II) sulfate monohydrate.
Clinical biochemistry findings:: no effects observed
Urinalysis findings:: not examined
Behaviour (functional findings):: not examined
Organ weight findings including organ / body weight ratios:: no effects observed
Description (incidence and severity):: no chemical related significant incidences over control in pancreas or kidney
Gross pathological findings:: no effects observed
Description (incidence and severity):: no chemical related significant incidences over control in pancreas or kidney
Histopathological findings: non-neoplastic:: no effects observed
Description (incidence and severity):: No incidences of non-neoplastic lesions were detected in both sexes of both species significantly over control
Histopathological findings: neoplastic:: no effects observed
Description (incidence and severity):: No incidences of neoplastic lesions were detected in both sexes of both species significantly over control
Details on results:: CLINICAL SIGNS AND MORTALITY
Rats: Survival of 15,000 ppm male rats was significantly lower than that of the controls; survival of 1,500 and 5,000 ppm males and all exposed groups of females was similar to that of controls. The significant reduction in survival of 15,000 ppm males was attributed to increased severity of nephropathy and renal failure. The decreased survival did not occur until approximately week 93 of the study.
Mice: Survival of exposed males and females was similar to that of the control groups. No clinical findings were attributed to the administration of manganese (II) sulfate monohydrate.

BODY WEIGHT AND WEIGHT GAIN
Rats: The mean body weights of 1,500 and 5,000 ppm male rats were similar to those of controls throughout the 2-year study. The mean body weights of 15,000 ppm male rats were within 5% of that of controls until week 89. From week 89, the mean body weights ranged from 8% to 13% lower than that of controls; at the end of the 2-year study, the final mean body weight of 15,000 ppm males was 10% lower than that of controls. Mean body weights of exposed females were similar to that of controls throughout the study.
Mice: After week 37, mean body weights of all exposed groups of females were lower than that of the controls; the final mean body weights for the 1,500, 5,000, and 15,000 ppm groups were 6%, 9%, and 13% lower than that of the control group.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study)
Rats: Feed consumption by exposed groups was similar to that by control groups.
Mice: Feed consumption by exposed male and female mice was similar to that of the control groups.

FOOD EFFICIENCY
No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study)
No data

OPHTHALMOSCOPIC EXAMINATION
No data

HAEMATOLOGY
Rats: Slight differences in some parameters between exposed and control groups were not considered related to the ingestion of manganese (II) sulfate monohydrate.
Mice: Slight differences in some parameters between exposed and control groups are found and not consistent with the findings in the 13-week study and their significance is uncertain.

CLINICAL CHEMISTRY
No data

URINALYSIS
No data

NEUROBEHAVIOUR
No data

ORGAN WEIGHTS
Rats: At the 9- and 15-month interim evaluations, the absolute kidney weights of exposed rats were similar to those of the controls.
Mice: At the 9-month interim evaluation, absolute liver weights of 15,000 ppm males and of 5,000 and 15,000 ppm females were significantly lower than those of controls. Since these groups also had lower mean body weights, and relative liver weights were similar to controls, the lower absolute liver weights are not considered chemical related. At the 15-month interim evaluation, absolute and relative liver weights of exposed mice were similar to controls.

GROSS PATHOLOGY
No data

HISTOPATHOLOGY: NON-NEOPLASTIC
Rats:
Pancreas: Hyperplasia or adenoma of the pancreatic islets occurred in a few males in each of the exposure groups but not in the control group. However, neither the trend test nor pairwise comparisons were significant, and the incidences in each of the dose groups were within the range of NTP historical control groups.
Kidney: Chronic nephropathy occurred in all male rats examined at both interim evaluations and most of the control and exposed males at the end of the study. The average severity of nephropathy was slightly greater in the high-dose group, but the difference was not statistically significant. Because of the subjective nature of the severity grading, an additional evaluation of the kidney of high-dose and control male rats was performed without knowledge of the previous diagnoses. The result of the additional evaluation confirmed the presence of a marginally increased severity of nephropathy in the high-dose group, and the difference was significant (P=0.04) by a two-sided MannWhitney U test. The severity of nephropathy varied from minimal to marked. Minimal nephropathy was characterized by a few sparsely scattered cortical foci of regenerating tubules with increased epithelial cytoplasmic basophilia and slightly thickened glomerular basement membranes. Nephropathy of mild severity had similar morphologic features, but these features occurred with greater frequency. Also present were occasional dilated tubules filled with homogenous hyaline material and lined by flattened epithelial cells. Nephropathy of moderate to marked severity had similar but more severe and extensive tubule lesions. In addition, variable interstitial fibrosis and mineralization with mononuclear leukocyte infiltration, variable tubule loss and atrophy, and degenerative glomerular changes occurred. In the most severe cases, cystic tubules lined by cuboidal or attenuated epithelial cells were present.
The incidences of several lesions commonly associated with advanced nephropathy and renal failure were significantly increased in 15,000 ppm male rats. These lesions included mineralization of blood vessels, mineralization of the glandular stomach, fibrous osteodystrophy of the femur, and parathyroid gland hyperplasia.
Mice:
Thyroid Gland: At the 9- and 15-month interim evaluations, thyroid follicle dilatation was present in 15,000 ppm males and females but not in the controls. At the end of the 2-year study, the incidence of follicular dilatation increased significantly in 15,000 ppm males and 5,000 and 15,000 ppm females. A significantly increased incidence of focal hyperplasia of follicular epithelium also occurred in 15,000 ppm males and in all exposed females. Follicular dilatation at the 9-month evaluation was characterized by a uniform increase in the follicular diameter throughout the gland. Follicular dilatation in mice at the end of the study differed from that observed in mice at the 9-month interim evaluation in that the dilated follicles were limited to the periphery of the glands. The affected follicles contained pale eosinophilic colloid and were lined by a single layer of flat to slightly cuboidal follicular epithelial cells. Follicular cell hyperplasia and adenoma constitute a morphological continuum. Follicular cell hyperplasia consisted of single or multiple collections of variably sized follicles with irregular hypertrophy and increased cellularity of the follicular epithelium. Minimal to mild follicular cell hyperplasia consisted of one or several follicles lined by columnar epithelium with small and infrequent papillary infoldings. Moderate to marked hyperplasia involved clusters of variably sized follicles with more prominent papillary formations.
Forestomach: A statistically significant increased incidence of focal squamous hyperplasia of the forestomach occurred in the 15,000 ppm males and females, accompanied by ulceration/erosion and inflammation. Hyperplasia of the squamous epithelium occurred focally at various sites of the forestomach mucosa. The lesion was characterized by broad-based areas of either proliferative epithelial thickening and hyperkeratosis or by polypoid projections of thickened epithelium protruding directly from the mucosa into the lumen of the stomach. Inflammation of the lamina propria and submucosa subjacent to the ulcerative lesions consisted of a mixture of infiltrating neutrophils and mononuclear leukocytes.

HISTOPATHOLOGY: NEOPLASTIC (if applicable)
Rats:
Pancreas: Hyperplasia or adenoma of the pancreatic islets occurred in a few males in each of the exposure groups but not in the control group. In addition, a carcinoma of the pancreatic islets was found in one 15,000 ppm male. However, neither the trend test nor pairwise comparisons were significant, and the incidences in each of the dose groups were within the range of NTP historical control groups.
Adrenal Gland: In females, medullary hyperplasia occurred with a significant negative trend and a significantly decreased incidence in the 15,000 ppm group. Benign pheochromocytomas of the adrenal medulla in males occurred with a significant negative trend, but the decreases were not significant by pairwise comparison the incidence of medullary hyperplasia in exposed males was similar to that of the controls
Mice:
Thyroid Gland: Follicular cell adenomas were found in three (6%) 15,000 ppm males. This rate is marginally higher than the average rate of 2% and just within the range of 0%-6% for historical control male mice. The incidence of this neoplasm was 10% in 15,000 ppm females, which is slightly above the average of 3% and range of 0%-9% for historical control female mice. The incidences of adenoma in 15,000 ppm males and females were not significantly greater than those of the controls. Follicular cell adenomas were generally more discrete collections of altered follicles compressing the surrounding parenchyma at the 9-month evaluation.
Liver: One male in the 15,000 ppm group and two females in the 5,000 ppm group had hepatocellular adenomas at the 15-month interim evaluation. At the end of the 2-year study, hepatocellular adenomas occurred with a statistically significant negative trend in males that was also significant by pairwise comparison in the 5,000 and 15,000 ppm groups. Hepatocellular foci did not occur in an exposure-related pattern. The incidences of adenoma or foci in exposed females were similar to those of the controls.

HISTORICAL CONTROL DATA (if applicable)
Lesion incidences from the NTP historical control database (Haseman et al., 1984, 1985) are included.

OTHER FINDINGS
Rats: At both the 9- and 15-month interim evaluations, the manganese levels in the liver of 5,000 and 15,000 ppm males and females were significantly greater than those in controls. The hepatic iron concentrations for these exposure groups were lower than for controls. The concentrations of manganese in the brain, kidney, and pancreas of exposed and control rats were variable; 15,000 ppm males had a significantly higher concentration of manganese in the brain and kidney at the 9-month interim evaluation and in the brain, kidney, and pancreas at the 15-month interim evaluation. Copper levels in the kidney of 15,000 ppm males at 9 months and in 15,000 ppm females at 9 and 15 months were significantly greater than those of the controls.
Mice: At the 9- and 15-month interim evaluations, tissue concentrations of manganese were significantly elevated in the livers of the 5,000 and 15,000 ppm groups. Hepatic iron levels were significantly lower in exposed females at the 9- and 15-month interim evaluations and in 5,000 and 15,000 males at the 15-month interim evaluation. Tissue concentrations of manganese in the brain (except 1,500 and 5,000 ppm females at 15 months), kidney, and pancreas (except 1,500 males at 9 months and 1,500 ppm females at 15 months) of exposed groups were significantly greater those of controls (Tables H3 and H4).

Effect levels

1

Dose descriptor:: NOEL
Effect level:: ca. 700 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: rat, mean survival, range 467-1171 mg/kg bw/day (highest dose tested)

2

Dose descriptor:: NOEL
Effect level:: ca. 790 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: rat, mean survival, range 561-1249 mg/kg bw/day (highest dose tested)

3

Dose descriptor:: NOEL
Effect level:: ca. 1 972 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: mouse, mean survival, range 1371-2663 mg/kg bw/day (highest dose tested)

4

Dose descriptor:: NOEL
Effect level:: ca. 2 500 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: mouse, mean survival, range 1636-3449 mg/kg bw/day (highest dose tested)

5

Dose descriptor:: NOAEL
Effect level:: ca. 700 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: rat, nephropathy, range 467-1171 mg/kg bw/day (highest dose tested)

6

Dose descriptor:: NOAEL
Effect level:: ca. 790 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: rat, nephropathy, range 561-1249 mg/kg bw/day (highest dose tested)

7

Dose descriptor:: NOAEL
Effect level:: ca. 600 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: Mouse, range 435-845 mg/kg bw/day Thyroid gland, Follicular dilatation and focal hyperplasia in follicular cells

8

Dose descriptor:: NOAEL
Effect level:: ca. 228 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: mouse, range 144-345 mg/kg bw/day Thyroid gland, Follicular dilatation

9

Dose descriptor:: NOAEL
Effect level:: ca. 600 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: Mouse, range 435-845 mg/kg bw/day Forestomach, focal squamous hyperplasia

10

Dose descriptor:: NOAEL
Effect level:: ca. 792 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: Mouse, range 526-1146 mg/kg bw/day Forestomach, focal squamous hyperplasia

11

Dose descriptor:: NOAEL
Effect level:: ca. 1 972 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: male
Basis for effect level:: other: mouse, relative liver weight, range 1371-2663 mg/kg bw/day (highest dose tested), at 15 month interim evaluation

12

Dose descriptor:: NOAEL
Effect level:: ca. 2 500 mg/kg bw/day (nominal)
Based on:: test mat.
Remarks:: MnSO4
Sex:: female
Basis for effect level:: other: mouse, relative liver weight, range 1636-3449 mg/kg bw/day (highest dose tested), at 15 month interim evaluation

Target system / organ toxicity

Critical effects observed:: not specified

Any other information on results incl. tables

TABLE 4 Survival of Rats in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

	Control	1,500 ppm	5,000 ppm	15,000 ppm
Male
Animals initially in study	70	70	70	70
9-Month interim evaluation^a	8	10	10	10
15-Month interim evaluation^a	10	9	9	8
Moribund	21	24	24	38
Natural deaths	6	10	5	7
Animals surviving to study termination	25	17	22^b	7
Percent probability of survival at end of study^cMean survival (days)^d	49	34	43	14
	581	573	579	571
Survival analyses^e	P=0.004	P=0.381	P=0.872	P=0.006
Female
Animals initially in study	70	70	70	70
9-Month interim evaluation^a	10	10	10	10
15-Month interim evaluation^a	10	10	9	10
Accidental deaths^a	0	0	1	0
Moribund	6	11	6	11
Natural deaths	7	2	2	1
Missexed^a	0	0	0	2
Animals surviving to study termination	37	37	42	36
Percent probability of survival at end of study	74	74	85	75
Mean survival (days)	608	594	596	607
Survival analyses	P=0.914N	P=0.986	P=0.378N	P=0.984N

a Censored from survival analyses

b Includes one animal that died the last week of study

c Kaplan-Meier determinations

d Mean of all deaths (uncensored, censored, and terminal sacrifice)

e The result of the life table trend test (Tarone, 1975) is in the control column, and the results of the life table pairwise comparisons (Cox, 1972) with the controls are in the exposed columns. A negative trend or lower mortality in an exposure group is indicated by N.

TABLE 5 Mean Body Weights and Survival of Male Rats in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Weeks on Study	0 ppm		1,500 ppm			5,000 ppm			15,000 ppm
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
1	126	70	124	98	70	126	100	70	123	98	70
2	167	70	166	100	70	167	101	70	164	98	70
3	201	70	197	98	70	202	101	70	193	96	70
4	227	70	220	97	70	228	100	70	220	97	70
5	247	70	244	98	70	251	101	70	244	99	70
6	272	70	268	98	70	274	101	70	268	98	70
7	283	70	280	99	70	287	102	70	280	99	70
8	297	70	295	99	70	303	102	70	293	99	70
9	314	70	307	98	70	313	100	70	306	98	70
10	327	70	318	97	70	328	100	70	321	98	70
11	334	70	327	98	70	334	100	70	324	97	70
12	343	70	335	98	70	343	100	70	335	98	70
13	353	70	347	98	70	351	99	70	341	97	70
18	386	69	375	97	70	383	99	70	376	97	70
21	399	69	394	99	69	394	99	70	386	97	70
25	415	69	408	98	69	411	99	70	399	96	70
28	431	69	423	98	69	425	98	70	416	96	70
32	447	69	440	98	69	441	99	70	429	96	70
36	462	69	455	99	69	453	98	70	442	96	70
40^a	460	60	453	98	60	452	98	60	441	96	58
44	472	60	450	95	60	451	96	60	441	93	58
48	480	60	474	99	58	476	99	60	466	97	58
53	484	60	479	99	57	479	99	60	471	97	57
57	496	60	487	98	57	487	98	59	482	97	57
61	497	60	485	98	57	490	99	58	481	97	57
65^a	505	50	488	97	48	493	98	49	488	97	49
69	509	49	487	96	46	491	97	49	486	95	49
73	510	48	493	97	46	497	97	47	493	97	47
77	509	47	490	96	45	496	98	43	490	96	46
81	498	46	479	96	45	484	97	42	477	96	45
85	492	43	479	97	44	484	99	41	478	97	44
89	486	38	469	97	40	469	97	39	449	92	43
93	472	35	451	96	38	460	97	35	432	91	35
97	449	31	426	95	32	443	99	32	409	91	26
101	433	26	431	100	23	423	98	28	378	87	18
104	402	26	402	100	20	393	98	24	360	90	10
Mean for weeks
1-13	269		264	98		270	100		262	97
14-52	439		430	98		432	98		422	96
53-104	482		468	97		471	98		455	94

a Interim evaluations occurred during weeks 39 and 65.

TABLE 6 Mean Body Weights and Survival of Female Rats in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Weeks on Study	0 ppm		1,500 ppm			5,000 ppm			15,000 ppm
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
1	104	70	103	100	70	104	100	70	103	99	70
2	127	70	128	101	70	128	101	70	126	99	69
3	143	70	143	100	70	143	100	70	142	99	69
4	154	70	154	100	70	153	99	70	151	98	68
5	161	70	164	102	70	160	99	70	160	99	68
6	170	70	173	102	70	170	100	70	168	99	68
7	174	70	179	103	70	176	101	70	175	101	68
8	181	70	184	102	70	181	100	69	181	100	68
9	186	70	190	102	70	186	100	69	186	100	68
10	190	70	194	102	70	190	100	69	190	100	68
11	191	70	195	102	70	192	100	69	191	100	68
12	196	70	198	101	70	196	100	68	195	100	68
13	199	70	201	101	70	199	100	68	198	100	68
18	205	70	208	101	70	206	100	68	205	100	68
21	210	70	213	101	70	211	100	68	211	100	68
25	217	70	221	102	70	218	100	68	217	100	68
28	225	70	225	100	70	222	99	68	223	99	68
32	232	70	236	102	70	231	100	68	231	100	68
36	237	70	244	103	70	240	101	68	237	100	68
40^a	241	60	245	102	60	240	101	58	243	101	58
44	249	60	257	103	60	251	101	58	251	101	58
48	258	60	260	101	60	261	101	58	266	103	58
53	272	60	277	102	60	270	99	58	277	102	58
57	277	60	287	104	60	279	101	58	287	104	58
61	286	60	294	103	59	287	101	58	295	103	58
65^a	296	50	305	103	48	298	101	49	303	103	48
69	305	50	313	103	48	306	101	49	312	102	48
73	311	50	320	103	47	316	101	49	318	102	47
77	321	50	327	102	47	328	102	48	329	103	46
81	327	49	332	102	43	333	102	48	335	103	46
85	335	48	339	101	42	340	102	47	342	102	46
89	336	47	346	103	41	346	103	45	346	103	46
93	340	43	342	101	41	345	102	45	345	102	45
97	334	43	335	100	39	341	102	43	337	101	44
101	336	38	330	98	39	337	100	42	333	99	38
104	327	38	326	100	37	336	103	42	336	103	36
Mean for weeks
1-13	167		170	102		168	101		167	100
14-52	230		234	102		231	100		232	101
53-104	315		320	102		319	101		321	102

a Interim evaluations occurred during weeks 39 and 65.

TABLE 7 Incidence and Severity of Nephropathy of Rats in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Dose (ppm)	0	1,500	5,000	15,000
Males
9-Month Interim Evaluation
Kidney^a	8	10	10	10
Nephropathy^b	8 (1.3)^c	10 (1.1)	10 (1.1)	10 (1.3)
15-Month Interim Evaluation
Kidney	10	9	9	8
Nephropathy	10 (1.6)	9 (1.9)	9 (1.9)	8 (2.0)
2-Year Study
Kidney	52	50	51	52
Nephropathy (initial evaluation)	50 (2.9)	49 (3.0)	51 (3.0)	50 (3.2)
Nephropathy (additional evaluation)	52 (2.8)	-	-	51 (3.1)*
Females
2-Year Study
Kidney	50	50	51	48
Nephropathy	48 (1.8)	50 (1.5)	49 (1.7)	48 (1.9)

* Significantly different (P0.05) from the control group by two-sided Mann-Whitney U test

a Number of animals with organ examined microscopically

b Number of animals with lesion

c Average severity grade of lesions in all animals (0=normal; 1=minimal; 2=mild; 3=moderate; 4=marked)

TABLE 10 Survival of Mice in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

	Control	1,500 ppm	5,000 ppm	15,000 ppm
Male
Animals initially in study	70	70	70	70
9-Month interim evaluation^a	10	10	10	9
15-Month interim evaluation^a	10	10	9	10
Accidental deaths^a	0	0	0	1
Moribund	2	3	2	1
Natural deaths	2	3	3	3
Animals surviving to study termination	46^b	44^b	46	46
Percent probability of survival at end of study^cMean survival (days)^d	92	88	91	93
	620	619	615	615
Survival analyses^e	P=0.920N	P=0.708	P=0.992	P=0.748
Female
Animals initially in study	70	70	70	70
9-Month interim evaluation^a	10	10	10	10
15-Month interim evaluation^a	9	10	9	9
Accidental deaths^a	1	0	0	0
Moribund	6	4	6	4
Natural deaths	2	0	6	5
Missing^a	0	0	1	0
Animals surviving to study termination	42	46	38	42
Percent probability of survival at end of study	85	92	77	83
Mean survival (days)	605	623	594	614
Survival analyses	P=0.563	P=0.318N	P=0.456	P=0.961

a Censored from survival analyses

b Includes one animal that died during the last week of the study.

c Kaplan-Meier determinations

d Mean of all deaths (uncensored, censored, and terminal sacrifice)

TABLE 11 Mean Body Weights and Survival of Male Mice in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Weeks on Study	0 ppm		1,500 ppm			5,000 ppm			15,000 ppm
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
1	21.1	70	21.1	100	70	21.1	100	70	20.6	98	70
2	22.9	70	22.7	99	70	22.8	100	70	22.1	97	70
3	23.8	70	23.9	100	70	23.6	99	70	23.1	97	70
4	24.7	70	25.1	102	70	24.8	100	70	24.2	98	70
5	25.6	70	26.1	102	70	25.7	100	70	25.0	98	70
6	26.2	70	26.9	103	70	26.4	101	70	25.8	99	70
7	27.4	70	28.1	103	70	27.6	101	70	26.9	98	70
8	27.7	70	28.4	103	70	27.6	100	70	26.9	97	70
9	28.7	70	29.2	102	70	28.8	100	70	27.8	97	70
10	29.1	70	29.4	101	70	29.1	100	70	28.1	97	70
11	29.9	70	30.3	101	70	30.1	101	70	29.0	97	70
12	30.5	70	31.3	103	70	30.9	101	70	29.5	97	70
13	31.6	70	32.5	103	70	31.8	101	70	30.6	97	70
17	34.1	70	35.2	103	70	34.6	102	70	32.8	96	70
21	36.9	70	37.9	103	70	37.1	101	70	35.4	96	69
25	39.0	70	40.1	103	70	39.3	101	70	37.6	96	69
29	41.4	70	42.1	102	70	41.4	100	70	39.3	95	69
33	43.3	70	43.7	101	70	43.1	100	70	41.1	95	69
37	43.8	70	44.5	102	70	43.9	100	69	42.1	96	69
41^a	44.5	60	45.2	102	60	44.8	101	59	43.2	97	60
45	45.4	60	46.5	102	60	46.0	101	59	44.4	98	60
49	46.8	60	48.2	103	60	47.4	101	59	45.8	98	60
54	46.7	60	48.0	103	60	47.3	101	59	46.0	99	60
57	46.0	60	47.6	103	60	46.5	101	59	45.7	99	60
61	46.6	60	48.0	103	60	47.1	101	58	46.4	100	60
65^a	47.0	50	48.4	103	50	47.3	100	49	46.4	99	49
69	46.9	49	48.2	103	50	47.1	100	49	45.7	97	49
73	46.8	49	48.1	103	50	46.5	99	49	45.3	97	49
78	46.4	49	48.3	104	50	47.1	102	49	46.0	99	49
82	47.7	49	49.3	103	50	48.2	101	49	46.6	98	49
86	47.8	49	48.9	102	49	48.2	101	48	46.8	98	49
90	48.4	49	48.9	101	48	48.7	101	48	47.4	98	49
94	47.0	49	47.6	101	47	47.0	100	48	46.2	98	46
98	46.6	49	48.1	103	46	47.0	101	48	45.7	98	46
101	45.9	48	46.9	102	44	44.9	98	47	44.0	96	46
104	45.2	47	46.3	102	44	44.9	99	47	44.1	98	46
Mean for weeks
1-13	26.9		27.3	102		26.9	100		26.1	97
14-52	41.7		42.6	102		42.0	101		40.2	96
53-104	46.8		48.0	103		47.0	100		45.9	98

a Interim evaluations occurred during weeks 39 and 65.

TABLE 12 Mean Body Weights and Survival of Female Mice in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Weeks on Study	0 ppm		1,500 ppm			5,000 ppm			15,000 ppm
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
	Av. Wt. (g)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors	Av. Wt. (g)	Wt. (% of controls)	No. of Survivors
1	17.2	70	17.0	99	70	17.1	99	70	17.0	99	70
2	18.4	70	18.4	100	70	18.5	101	70	18.0	98	70
3	19.5	70	19.5	100	70	19.5	100	70	19.2	99	70
4	20.6	70	20.7	101	70	20.7	101	69	20.0	97	70
5	21.6	70	21.7	101	70	21.5	100	69	20.7	96	70
6	21.9	70	22.0	101	70	21.9	100	69	21.3	97	70
7	23.0	70	22.8	99	70	22.9	100	69	22.3	97	70
8	23.4	70	23.8	102	70	23.6	101	69	22.9	98	70
9	24.3	70	24.3	100	70	24.3	100	69	23.9	98	70
10	24.5	70	24.4	100	70	24.4	100	69	23.8	97	70
11	25.1	70	25.1	100	70	25.1	100	69	24.6	98	70
12	25.6	70	25.6	100	70	25.7	100	69	25.2	98	70
13	25.6	70	26.2	102	70	26.3	103	69	25.5	100	70
17	28.1	70	28.8	103	70	28.6	102	68	27.7	99	70
21	30.4	70	31.4	103	70	31.5	104	68	30.5	100	70
25	33.0	70	33.7	102	70	33.9	103	68	32.8	99	70
29	36.0	70	35.7	99	70	36.0	100	68	34.5	96	70
33	38.5	70	38.3	100	70	38.2	99	68	37.0	96	70
37	40.3	70	39.7	99	70	39.3	98	68	37.9	94	70
41^a	41.9	59	41.6	99	60	40.9	98	58	39.0	93	60
45	43.8	59	43.6	100	60	42.7	98	58	41.6	95	60
49	46.3	58	46.6	101	60	45.4	98	58	43.6	94	60
54	47.6	58	47.0	99	60	45.8	96	58	43.8	92	60
57	47.6	57	47.4	100	60	46.0	97	58	43.9	92	60
61	48.6	57	48.4	100	60	46.6	96	57	45.0	93	60
65^a	49.7	48	48.6	98	50	47.0	95	48	45.3	91	50
69	49.8	48	49.5	99	50	47.8	96	48	45.6	92	49
73	51.0	48	50.7	99	50	48.1	94	47	46.3	91	48
78	50.6	47	50.5	100	50	48.4	96	47	46.3	92	48
82	52.7	47	51.6	98	50	50.1	95	46	47.9	91	48
86	53.1	47	52.0	98	50	50.0	94	46	47.8	90	48
90	53.5	45	51.8	97	50	50.8	95	44	48.3	90	48
94	53.8	45	51.0	95	50	50.5	94	44	47.7	89	47
98	52.4	45	49.7	95	49	48.6	93	44	46.2	88	46
102	51.6	43	48.4	94	48	47.2	92	40	44.4	86	44
104	50.8	42	47.8	94	46	46.2	91	40	44.3	87	43
Mean for weeks
1-13	22.4		22.4	100		22.4	100		21.9	98
14-52	37.6		37.7	100		37.4	99		36.1	96
53-104	50.9		49.6	97		48.1	94		45.9	90

a Interim evaluations occurred during weeks 39 and 65.

TABLE 13 Incidences of Selected Lesions of the Thyroid Gland of Mice in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Dose (ppm)	0	1,500	5,000	15,000
Males
9-Month Interim Evaluation
Thyroid Gland^a	10	0	0	9
Follicular Dilatation^b	0	^_c	-	6**(1.0)^d
15-Month Interim Evaluation
Thyroid Gland	10	2	1	10
Follicular Dilatation	0	0	0	9**(1.0)
Follicular Cell Adenoma	0	0	0	1
2-Year Study
Thyroid Gland	50	49	51	50
Follicular Dilatation	2 (1.0)	2 (1.5)	5 (1.0)	23** (1.2)
Follicular Cell, Hyperplasia, Focal	5 (1.0)	2 (1.5)	8 (1.5)	27** (1.9)
Follicular Cell Adenoma^e
Overall rate^f	0/50 (0%)	0/49 (0%)	0/51 (0%)	3/50 (6%)
Adjusted rate^g	0.0%	0.0%	0.0%	6.5%
Terminal rate^h	0/46 (0%)	0/44 (0%)	0/46 (0%)	3/46 (7%)
First incidence (days)	-	-	-	729 (T)
Logistic regression testⁱ	P=0.015	-	-	P=0.121
Females
9-Month Interim Evaluation
Thyroid Gland	10	0	1	10
Follicular Dilatation	0	-	1 (1.0)	7** (1.0)
15-Month Interim Evaluation
Thyroid Gland	9	0	2	9
Follicular Dilatation	0		0	5* (1.0)
2-Year Study
Thyroid Gland	50	50	49	51
Follicular Dilatation	1 (1.0)	5 (1.0)	11** (1.4)	24** (1.2)
Follicular Cell, Hyperplasia, Diffuse	1 (1.0)	1 (1.0)	0	0
Follicular Cell, Hyperplasia, Focal	3 (2.3)	15** (1.5)	27** (1.5)	43** (2.1)
Follicular Cell Adenoma^j
Overall rates	2/50 (4%)	1/50 (2%)	0/49 (0%)	5/51 (10%)
Adjusted rates	4.8%	2.2%	0.0%	11.9%
Terminal rates	2/42 (5%)	1/46 (2%)	0/37 (0%)	5/42 (12%)
First incidence (days)	729 (T)	729 (T)		729 (T)
Logistic regression tests	P=0.037	P=0.468N	P=0.267N	P=0.216

(T)Terminal Sacrifice

* Significantly different (P0.05) from the control group by the Fisher exact test (interim evaluations) or by the logistic regression test (2-year study)

** P≤0.01

a Number of animals with organ examined microscopically

b Number of animals with lesion

c Not applicable; tissue not examined microscopically in this group

d Average severity grade of lesions in affected animals (1=minimal; 2=mild; 3=moderate; 4=marked)

e Historical incidence for 2-year feed studies with untreated control groups (mean ± standard deviation): 19/1,105 (1.7% ± 1.7%); range 0%-4%

f Number of neoplasm-bearing animals/number of animals microscopically examined

g Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality

h Observed incidence at terminal kill

i Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to pairwise comparisons between the controls and that exposed group. The logistic regression test regards these neoplasms as nonfatal. A lower incidence in an exposure group is indicated by N.

j Historical incidence: 27/1,099 (2.5% ± 2.9%); range 0%-9%

TABLE 14 Incidences of Selected Lesions of the Forestomach of Mice in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate

Dose (ppm)	0	1,500	5,000	15,000
Males
2-Year Study
Forestomach^a	50	49	51	50
Erosion, Focal^b	0	0	0	2 (3.0)^c
Squamous Hyperplasia, Focal	2 (2.0)	1 (2.0)	5 (1.2)	14** (2.3)
Inflammation, Chronic Active	0	0	0	5* (2.0)
Ulcer	0	0	0	6* (2.5)
Squamous Cell Papilloma	1	1	0	0
Females
9-Month Interim Evaluation
Forestomach	10	0	0	9
Squamous Hyperplasia, Focal	0	0	0	1 (2.0)
15-Month Interim Evaluation
Forestomach	9	0	1	9
Squamous Hyperplasia, Focal	0	0	1 (2.0)	1 (2.0)
Inflammation, Chronic Active	0	0	0	1 (1.0)
2-Year Study
Forestomach	51	50	49	50
Squamous Hyperplasia, Focal	1 (2.0)	3 (1.7)	3 (2.0)	9**(2.4)
Ulcer	2 (2.0)	0	0
3 (2.7)
Inflammation, Chronic Active	0	1 (2.0)	1 (2.0)	3 (1.7)
Squamous Cell Papilloma	1	0	0	0

* Significantly different (P0.05) from the control group by the logistic regression test

** P0.01

a Number of animals with organ examined microscopically

b Number of animals with lesion

c Average severity grade of lesions in affected animals (1=minimal; 2=mild; 3=moderate; 4=marked)

TABLE A1 Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of Manganese (II) Sulfate Monohydrate^a