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Administrative data

Endpoint:
health surveillance data
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: A well documented study conducted to good scientific principles.

Data source

Reference
Reference Type:
publication
Title:
Intestinal effects of sulfate in drinking water on normal human subjects
Author:
Heizer W D, Sandler R S, Seal E, Murray SC, Busby M, Schliebe BG & Pusek SN.
Year:
1997
Bibliographic source:
Digestive Diseases and Science 42 (5) 1055 - 1061

Materials and methods

Study type:
self-reporting of symptoms
Endpoint addressed:
basic toxicokinetics
repeated dose toxicity: oral
Test guideline
Qualifier:
no guideline available
Principles of method if other than guideline:
A controlled study was conducted in normal adults to determine the effect of various drinking water sodium sulfate concentrations on bowel function. Ten healthy subjects were given a constant diet and fluid intake. Fluid consisted of 36 mL/kg/day of drinking water of various known sulfate concentration and 500 mL of other fluid. In a dose ranging study, four subjects received drinking water with sulfate concentratons of 0, 400, 600, 800, 1000 and 1200 mg/L for six consecutive two day periods. In a single dose study, six other subjects received water with sulfate concentrations of 0 and 1200 mg/L for two two consecutive six-day periods. Stool mass, frequency, and consistency and mouth-to-anus appearance time of coloured markers were measured.
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material: anhydrous sodium sulfate
- Analytical purity: USP verified

Method

Type of population:
general
Ethical approval:
confirmed and informed consent free of coercion received
Remarks:
Subjects volunteered after being fully informed
Details on study design:
Ten subjects aged 24 - 45 years old, 50 % male and 80 % Caucasian were studied. All were in good health as determined by physical examination, history, urinalysis, blood cell counts and serum chemistries. The women had negative blood pregnancy tests. Subjects were excluded if they had a history of having more than three bowel movements per day or fewer than three per week, had used antibiotics or tobacco within two months, habitually ingested more than 5 cups of coffee daily, or abused alcohol or drugs. All were within 20 % of desirable weight. As bowel habits tend to vary with menstrual cycle, studies were begun in females 5-days after completion of a menstrual period. Subjects volunteered after being fully informed. All 10 completed the study which was approved by the University of North Carolina School of Medicine Committee on the Protection of the Rights of Human Subjects.

Subjects were admitted to the General Clinical Research Centre at 7 am on Day 1 of the study. Diets were the same composition for each subject, but the amounts were calculated to match each individual's usual daily caloric intake. Each subject received two different daily menus given on alternate days so that over each two-day period a subject's diet was constant. Sulfur contents of the two menus, estimated from available tables were 0.51 and 0.47 mmol/kg body weight/day (mean 0.49 mmol/kg/day). The amount of tea and coffee were also constant each day. Subjects ingested 36 mL/kg/day of drinking water including tea and coffee made from drinking water. The rate of ingestion was controlled at 6 mL/kg with each meal and 6 mL/kg between meals. This level of water intake was chosen because it is approximately two standard-deviations above the mean daily consumption for men and women based on interviews with 645 individuals by a trained nutritionist using a validated food frequency questionnaire. Fluids other than drinking water were limited to 500 mL/day and were the same each day. Subjects were required to ingest all of the food and water offered and they ate and drank only what was prepared by the General Clinical Research Centre, except in some cases when bag lunches were provided.
Most subjects spent the night in the General Clinical Research Centre and those who did not took drinking-water and urine collection containers with them each night and agreed to comply with the requirements of the study.

Stool and urine measurements:
Coloured markers given orally at the beginning of each change in drinking water sulfate concentration consisted of capsules containing 250 mg carmine red alternating with capsules of FD&C green food dye. All stools were collected individually in pre-weighed plastic containers, net weight determined and the colour recorded. Stools beginning with the first stool containing one marker and ending with the last stool before appearance of the next marker constituted a pool representing food and water intake between the time the two markers were ingested. The markers also permitted a measure of mouth-to-anus appearance time associated with each drinking water sulfate concentration. A dye marker was often observed in more than one stool and when markers were given 48 h apart in the dose-ranging study, the stool was sometimes still coloured by one marker when the next one appeared. The consistency of each stool was recorded by the subject on a 5-point scale as detailed below. Each stool was also rated by a nurse. On the few occasions when the patient and nurse ratings disagreed, a final rating was determined by the study coordinator. Throughout the study, each subject kept a diary of gastrointestinal symptoms, including abdominal cramping, bloating and diarrhoea. Twenty-four hours urine was collected daily in plastic containers and maintained at 4ºC. Aliquots were analyzed for inorganic sulfate using a turbidometric assay after precipitation of sulfate with barium.

Two men and two women participated in a dose-ranging study. Each received drinking water with sulfate concentrations of 0, 400, 600, 800, 1000 and 1200 mg/L (0 - 12.5 mmol/L) in the order listed for six consecutive two-day periods. Six additional subjects, three men and three women received drinking water with sulfate concentrations of 0 and 1200 mg/L for two consecutive 6-day periods. Subjects were not specifically informed of the sulfate concentrations in the water ingested at various stages of the study but were able to discern some differences in taste.

Drinking water preparation and analysis:
Distilled, deionized water and anhydrous sodium sulfate, USP, were used to prepare drinking water ranging in sulfate concentration from 0 to 1200 mg/L. The volume required each day for a subject plus 125 mL was prepared and an aliquot saved in a plastic container for analysis. Sulfate concentration in the distilled water was less than 1.0 mg/L. A sample of drinking water from each persons dwelling place was also analysed for sulfate. Drinking water analyses were performed according to the Methods for Chemical Analysis of Water and Waste, EPA Manual 600/4-79-020.

The Wilcoxon signed-rank test was used to compare the effects of specific concentrations of sulfate in drinking water with the effect of distilled water. Page's test was used to evaluate trends

Results and discussion

Results:
Dose-ranging study:
The subjects reported no diarrhoea in their daily diaries. If diarrhoea is defined as a passage of more than 250 g of stool per 24 h, then one subject had diarrhoea on four days; while ingesting water with 0, 600, 800 and 1000 mg/L, one subject had one day of diarrhoea on 1000 mg sulfate/L and two subjects each had one day of diarrhoea on 1200 mg/L. No subject passed more than 3 stools on any one day. The only complaint in the daily diaries of the four subjects was mild abdominal cramps reported by one subject for 2 days while receiving distilled water.

Single-dose study:
Compared to distilled water, water containing sulfate at 1200 mg/L produced a statistically significant increase in the mean stool mass per six-day pool, from 621 g to 922 g (P = 0.03) and in mean stool mass per hour from 4.8 g to 6.6 g (P = 0.03). Each subject showed an increase in stool mass per pool and in stool mass per hour. The mean 24-h (8 AM to 8 AM) stool mass was 103 g/day on distilled water and 156 g/day when drinking water containing 1200 mg/L. Stool frequency, stool consistency and mouth-to-anus appearance time were not significantly different on the high-sulfate water. Two of the six subjects reported abdominal cramps on the first day of starting the sulfate containing water. No other symptoms were reported.
None of the subjects reported diarrhoea as a symptom in the daily diary. Defining diarrhoea as stool mass greater than 250 g/day, three subjects did not experience diarrhoea at any time during the study. The remaining three subjects experienced an increased number of days with stool weight >250 g/day when their drinking water contained 1200 mg sulfate/L. However, for all six subjects the difference between the two dosage levels in terms of number of days with stool > 250 g/day was not significant using the Wilcoxon signed rank test (P = 0.25).

All blood and urine test results remained normal at the end of the investigation.
In retrospect, eight subjects rated the taste of water containing 1200 mg sulfate/L as neutral to slightly unpleasant, and one rated it very unpleasant. Mean sulfate concentration in the usual drinking water for the subjects was 17.9 mg/L (0.186 mmol/L) range 1.4 - 32.9 mg/L (0.015 - 0.343 mmol/L).
Dose-ranging study:
Measured sulfate concentrations differed from the nominal values by a mean of -1.6 % (range -7.5 % to +2.7 %). At the highest concentration (1200 mg/L) the mean total daily sulfate intake in drinking water for the 4 subjects was 2679 mg (27.9 mmol).
Increasing the sulfate concentration in drinking water every 48 h from 0 to 1200 mg/L produced no significant trend in stool mass per pool or mass per hour, based on Page's L-statistic. During the six periods when drinking water sulfate was 0, 400, 600, 800, 1000 and 1200 mg/L, the mean number of stools per pool were 2.5, 3.0, 2.3, 3.0, 2.0 and 2.8 respectively and the mean consistency ratings were 3.5, 3.3, 3.1, 3.4, 3.0 and 2.7, respectively, neither of which is a significant trend. There was a significant trend towards decreasing mouth-to-anus appearance time with increasing sulfate concentration (p = 0.03); the mean appearance time (h) were were 27.3, 17.9, 26.0, 16.1, 19.2 and 17.2 respectively.
There were no significant differences in any of the measurements between ingestion of 0 and 1200 mg/L as determined by the Wilcoxon signed rank test (P> 0.05).

Single-dose study:
The water prepared to contain sulfate at 1200 mg/L differed from the nominal value by a mean of +1.4 % (range of -3.3 % to +6.8 %). Mean total daily sulfate intake in drinking water at the 1200 mg/L concentration was 2908 mg (range 2256 - 3560 mg) or a mean of 30.3 mmol/day.
The mean stool mass (g/24 h) for days 1 - 6 of high sulfate intake was 125, 72, 230, 171, 175 and 156, respectively. This does not indicate a trend toward decreasing stool mass and therefore does not suggest that subjects were developing a tolerance to the effects of sulfate on stool mass.

When all 10 subjects in both the dose-ranging and single-dose studies were used to compare effects at 0 and 1200 mg/L, significant decreases in stool consistency (P = 0.02) and appearance time (P = 0.03) were noted at 1200 mg/L. An increase in stool mass per hour did not reach significance (P = 0.06). Stool mass per pool could not be compared as pools represented two days in the four subjects in the dose-ranging study and six-days in the six subjects in the single dose study.

Estimated absorption of sulfate in drinking water:
There was no significant trend toward decreasing percent absorption with increasing concentrations of sulfate in drinking water over the range tested in this study. A mean of 64 % of the sulfate ingested in drinking water was absorbed and excreted in the nine evaluable subjects (range 40 - 81 %).

Urinary sulfate derived from the diet:
During baseline periods when the drinking water contained no sulfate, mean urinary excretion of inorganic sulfate was 0.29 mmol/kg/day. Mean dietary sulfur was 0.49 mmol/kg/day. The finding that urinary excretion accounts for approximately 60 % of dietary sulfate in part results from the fact that approximately 20 % of urinary sulfate is ordinarily bound to organic compounds and would not be measured in the assay used.

Any other information on results incl. tables

Effect of sulfate in drinking water on stool mass: dose-ranging study (g/pool)

Subject

Stool mass* at g/pool†

0

400

600

800

1000

1200

1

308

421

343

476

278

372

2

420

363

396

332

339

357

3

286

296

98

380

230

287

4

48

306

92

119

201

382

Mean

265

347

23

327

262

350

SD

156

58

160

151

60

43

* There is no significant trend in stool mass with increasing sulfate concentration by Page's L statistic.

Sum of stool weights beginning with the first stool containing a marker and ending with the stool preceding the stool containing the next marker. Markers were given orally 48 h apart between 6:30 AM and 8:00 AM.

Effects of sulfate in drinking water on stool mass: dose-ranging study (g/hr)

Subject

Stool mass* at g/hr

0

400

600

800

1000

1200

1

7

12.1

8

7.4

8.9

8.5

2

8.2

8.1

8

7.2

7

10

3

10

3.4

9.1

5.3

9.4

3.2

4

6.6

4.7

7.4

1.8

3

13

Mean

8

7.1

8.1

5.4

7.1

9

SD

1.5

3.9

0.7

2.6

2.9

4.5

* There is no significant trend in stool mass with increasing sulfate concentration by Page's L ststistic.

‡ Pool weight divided by the hours during which the pool was collected measured from the times of the stools preceding the stools containing two consecutive markers. The mean pool duration was 46.3 hr.

Effects of sulfate in drinking water on stool mass: single dose study ( g/6-day pool)

Subject

Stool wt (g/6-day pool)*

0 mg sulfate/L

1200 mg sulfate/L

5

506

726

6

606

1083

7

197

424

8

932

1103

9

409

673

10

1073

1524

Mean

621

922‡

SD

328

392

* as for above tables. Markers were given at 6 -day (144-hr) intervals.

‡ Different from results at 0 sulfate/L by the Wilcoxon signed rank test (P = 0.03).

Effects of sulfate in drinking water on stool mass: single dose study ( g/h)

Subject

Stool output (g/hr)*

0 mg sulfate/L

1200 mg sulfate/L

5

3.6

5.8

6

4.7

7.9

7

1.9

2.5

8

7.4

8.6

9

3.3

4.6

10

8.1

10.4

Mean

4.8

6.6‡

SD

2.4

2.9

* as for above tables. Markers were given at 6-day (144-hr) intervals.

‡ Different from results at 0 sulfate/L by the Wilcoxon signed rank test (P = 0.03).

Effects of sulfate in drinking water on stool mass: single dose study (no. of days >250 g stool)

Subject

Number of days > 250 g stool

0 mg sulfate/L

1200 mg sulfate/L

5

0

0

6

1

2

7

0

0

8

1

2

9

0

0

10

1

3

Mean

0.5

1.2

SD

0.55

1.3

† number of days in which stool mass >250 g out of the total of 6 days for each sulfate concentration.

Urinary excretion of sulfate during dose-ranging study:

Subject

Percent of dose excreted*

Baseline excretion (mg/day)

Drinking water sulfate (mg/L)

Excretion (mean %)

400

600

800

1000

1200

1

2209

74.5

67.0

85.5

80.1

88.9

79.2

2

2450

0

23.0

0

32.0

9.0

-

3

1993

76.0

90.0

87.0

72.2

81.0

81.3

4

1345

44.0

78.0

80.0

84.0

60.0

69.2

Mean†

1999

64.8

78.3

84.2

78.9

76.6

76.6

SD

474

18.1

11.5

3.6

5.7

14.9

 

* there is no significant trend to decreasing percent of dose excreted with increasing sulfate concentration using Page's L statistic.

† subject 2 omitted from calculation of percentage dose excreted, but not from calculation of baseline excretion.

Urinary excretion of sulfate during single-dose study:

Subject

MeanBaseline

excretion

(mg/day)*

Fraction of dose

excreted

 (%)†

5

1993

40.1

6

1393

39.6

7

1273

79.8

8

2025

69.9

9

1305

57.2

10

2025

55.8

Mean

1669

57.1

SD

381

16.0

* mean of 6 days when drinking water contained no sulfate

† mean excretion of the sulfate ingested in drinking water (1200 mg/L).

 

Applicant's summary and conclusion

Conclusions:
In normal adult subjects, sulfate in drinking water at a concentration of 1200 mg/L, which is higher than reported to occur in US municipal water sources, caused a measureable but clinically insignificant increase in stool mass and decrease in stool consistency and appearance time, but no change in stool frequency and no complaint of diarhhoea.