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Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
supporting study
Reliability:
3 (not reliable)
Rationale for reliability incl. deficiencies:
other: Methodological deficiencies.

Data source

Reference
Reference Type:
publication
Title:
Greater effect of dietary potassium tripolyphosphate than of potassium dihydrogenphosphate on the nephrocalcinosis and proximal tubular function in female rats from the intake of a high-phosphorus diet.
Author:
Matsuzaki H, Masuyama R, Uehara M, Nakamura K, Suzuki K
Year:
2001
Bibliographic source:
Biosci Biotechnol Biochem. 65(4):928-34; PMID: 11388474

Materials and methods

Objective of study:
absorption
distribution
excretion
Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Rats were fed on a diet containing potassium dihydrogenorthophosphate or pentapotassium triphosphate at the normal phosphorus level or at a high phosphorus level for 21 d.
Faeces and urine were collected on days 17-20 by use of metabolic cages. After the treatment, the animals were sacrificed, blood was obtained by exsanguination and the kidneys were removed.
One kidney, feces and urine samples were ashed and analyzed for calcium, magnesium and phosphorus. Further urinalysis was also conducted. The other kidney was examined histopathologically and stained with Von Kossa stain to detect renal calcification.
GLP compliance:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): potassium dihydrogenphosphate, KH2PO4
- Storage condition of test material: 4°C
Radiolabelling:
no

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Clea Japan, Tokyo, Japan
- Age at study initiation: four weeks old
- Weight at study initiation: 102-104 g
- Housing: individually in stainless steel wire-mesh cages, on day 17-20: metabolic cages
- Diet (e.g. ad libitum): based on the AIN-93G diet ad libitum
- Water (e.g. ad libitum): demineralised water ad libitum
- Acclimation period: one week with a diet containing 100 mmol phosphorus /kg diet.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±1°C
- Humidity (%): 60-65%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: feed
Vehicle:
unchanged (no vehicle)
Details on exposure:
DIET PREPARATION
- Mixing appropriate amounts with (Type of food): standard diet (based on AIN-93G)
- Storage temperature of food: 4°C
Duration and frequency of treatment / exposure:
21 days, daily exposure
Doses / concentrations
Remarks:
Doses / Concentrations:
normal-phosphate diet: 100 mmol phosphorus per kg diet, equivalent to 640 mg/kg bw/day (assuming an average rat weight of 140 g)
high-phosphate diet: 400 mmol phosphorus per kg diet, equivalent to 4404 mg/kg bw/day (assuming an average rat weight of 140 g)
No. of animals per sex per dose:
6 female rats per dose
Control animals:
no
Details on study design:
- Rationale for animal assignment (if not random): similar mean body weight in the groups
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled (delete / add / specify): urine, faeces, blood, kidneys
- Time and frequency of sampling: days 17-20

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled (delete / add / specify): urine, faeces, kidney tissues
- Time and frequency of sampling: days 17-21
- From how many animals: mean of all 6 animals per group
- Method type(s) for identification: AAS
- Limits of detection and quantification: not given
Statistics:
The apparent absorption of minerals was calculated as the intake - faecal excretion, and the rate of apparent absorption as (intake - fecal excretion)/intake x 100. Each data value is expressed as the mean ± SE. Data were analyzed by two-way ANOVA to determine the effect of the dietary phosphorus level and the form of the phosphate salt. Tukey's test was used to determine the significance of differences in multiple comparisons among groups, differences being considered significant at P < 0.05. All statistical analyses were performed with the SPSS package program ver. 6.15.

Results and discussion

Toxicokinetic / pharmacokinetic studies

Details on absorption:
The absorption of calcium, magnesium and phosphate was calculated by subtraction of fecal excretion values by these orally administered.
In the high-dose group, the oral absorption of phosphate was significantly increased compared to the normal-phosphate diet whereas the uptake of calcium and magnesium were diminished. The values are depicted in table 6.
Details on distribution in tissues:
Concentrations of calcium, magnesium and phosphorus were non-significantly increased in the animals fed the high-phosphorus diet. For calcium the increase was not significant although the amount of calcium in the kidneys of the high-phosphorus group was increased by factor 14 compared to the normal-phosphorus diet animals. The relative amount was increased by factor 1.2 for both magnesium and phosphorus. The values are depicted in table 4.
Details on excretion:
Urinary phosphorus level was significantly increased and calcium and magnesium levels were significantly decreased by high-phosphate diet (see table 5).

Metabolite characterisation studies

Metabolites identified:
not measured

Any other information on results incl. tables

Table 3: Body weight and intake of food and minerals in rats fed on a diet containing different amounts of phosphate salts.

 

Normal-phosphorus diet

High-phosphorus diet

Phosphate amount

KH2PO4 [g/kg diet]

6.848

 

46.361

 

KH2PO4 [mmol/kg diet]

100

 

397

 

Body weight

           Initial [g]

102 ± 2

 

103 ± 1

 

           Final [g]

173 ± 2

 

175 ± 1

 

Food intake

Food [g/d]

13.1 ± 0.2

 

13.3 ± 0.3

 

Calcium [mmol/d]

1.88 ± 0.06

 

1.81 ± 0.03

 

Magnesium [mmol/d]

0.299 ± 0.008

 

0.272 ± 0.004

*

Phosphorus [mmol/d]

1.39 ± 0.04

 

5.42 ± 0.08

*

Each value is the mean ± SE, n=6 per group.

* Values are significantly different (P<0.05).

 

Table 4: Kidney analysis for rats fed on a diet containing different amounts of phosphate salts.

 

Normal-phosphorus diet

High-phosphorus diet

Dry weight [g/100g bw]

0.089 ± 0.001

 

0.099 ± 0.004

 

Calcium [mmol/100g dry weight]

1.01 ± 0.03

 

14.1 ± 2.0

 

Magnesium [mmol/100g dry weight]

3.62 ± 0.06

 

4.38 ± 0.16

 

Phosphorus [mmol/100g dry weight]

42.1 ± 1.4

 

50.4 ± 1.5

 

Severity of nephrocalcinosis

Score 0

6/6

0/6

Score 1

0/6

3/6

Score 2

0/6

3/6

Score 3

0/6

0/6

Each value is the mean ± SE, n=6 per group.

* Values with different letters in the same row are significantly different.

Score for nephrocalcinosis: 0 (not detected) < 1 < 2 < 3 (severe)

 

Table 5: Indicators of kidney function in rats fed on a diet containing different amounts of phosphate salts.

 

Normal-phosphorus diet

High-phosphorus diet

In blood

Urea nitrogen in serum [mmol/L]

9.53 ± 0.21

9.04 ± 0.43

In urine

Creatinine [µmol/d]

39.1 ± 1.4

 

41.5 ± 2.0

 

Albumin [g/mol creatinine]

2.60 ± 0.19

 

4.23 ± 0.87

 

NAG activity [U/mmol creatinine]

1.98± 0.21

 

2.51± 0.15

 

beta-Microglobulin [g/mol creatinine]

0.046± 0.004

 

0.060± 0.005

 

Calcium [mol/mol creatinine]

0.862± 0.065

 

0.251± 0.039

*

Magnesium [mol/mol creatinine]

3.03± 0.12

 

1.88± 0.10

*

Phosphorus [mol/mol creatinine]

3.03± 0.26

 

71.3± 3.2

*

Each value is the mean ± SE, n=6 per group.

* Values with different letters in the same row are significantly different.

 

Table 6: Apparent absorption of minerals by rats fed on a diet containing different amounts of phosphate salts.

 

 

Normal-phosphorus diet

High-phosphorus diet

 

Calcium

[mmol/d]

0.993 ± 0.038

0.898 ± 0.040

 

[%]

52.9 ± 1.7

49.6 ± 2.3

 

Magnesium

[mmol/d]

0.206 ± 0.006

0.119 ± 0.005

*

[%]

69.0 ± 1.4

43.9 ± 1.9

*

Phosphorus

[mmol/d]

0.817 ± 0.031

4.55 ± 0.09

*

[%]

58.9 ± 1.5

83.9 ±0.8

*

Each value is the mean ± SE, n=6 per group.

* Values with different letters in the same row are significantly different.

 

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): bioaccumulation potential cannot be judged based on study results
Administration of a diet with increased levels of potassium dihydrogenorthophosphate (4404 mg/kg bw/day) led to increased phosphate absorption compared to a diet containing "normal" levels of potassium dihydrogenorthophosphate (640 mg/kg bw/day). This phosphate was incorporated in the kidney resulting in marked nephrocalcinosis affecting renal function. Correspondingly, the amount of phosphate excreted in urine was significantly elevated.
Executive summary:

To assess whether the amount of daily administered dietary potassium dihydrogenorthophosphate influences nephrocalcinosis, this compound was administered for 21 days to female Wistar. The rats received either a normal-phosphate diet containing 640 mg/kg bw/day or a high-phosphate diet (4404 mg/kg bw/day). Faeces and urine were collected on days 17-20 by use of metabolic cages. After the treatment, the animals were sacrificed, blood was obtained by exsanguination and the kidneys were removed. One kidney, faeces and urine samples were ashed and analyzed for calcium, magnesium and phosphorus. Further urinalysis was also conducted. The other kidney was examined histopathologically and stained with Von Kossa stain to detect renal calcification.

Administration of a diet with increased levels of potassium dihydrogenorthophosphate led to increased phosphate absorption compared to a diet containing "normal" levels of potassium dihydrogenorthophosphate. This phosphate was incorporated in the kidney resulting in marked nephrocalcinosis affecting renal function. Correspondingly, the amount of phosphate excreted in urine was significantly elevated.