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Specific investigations: other studies

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Administrative data

Type of information:
experimental study
Adequacy of study:
supporting study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Limited description of experimental details.

Data source

Reference Type:
Determination of Spatial Distribution of Melamine-Cyanuric Acid Crystals in Rat Kidney Tissue by Histology and Imaging Matrix-Assisted Laser Desorption/Ionization Quadrupole Time-of-Flight Mass Spectrometry
Kim CW, Yun J-W, Bae I-H, Lee J-S, Kang H-J, Joo K-M, Jeong H-J, Chung J-H, Park Y-H,† and Lim K-M
Bibliographic source:
Chem. Res. Toxicol. 2010, 23, 220–227

Materials and methods

Principles of method if other than guideline:
Spatial distribution of melamine cyanurate crystals in kidney after 3-day feed application
GLP compliance:
Type of method:
in vivo
Endpoint addressed:
repeated dose toxicity: oral

Test material

Constituent 1
Details on test material:
obtained from Sigma- Aldrich (St. Louis, MO, USA).

Test animals

Details on test animals or test system and environmental conditions:
age: 5-6 months old
body weight: 250-270 g
Supplier: Orientbio, Seoul, Korea

The animals were housed in polyethylene cages in an environment with a controlled temperature of 24 ( 2 °C, a constant humidity of 50 ( 10%, and a 12 h light-dark cycle. Animals had free access to a standard diet from Purina Korea and tap water ad libitum. The rats were acclimatized for at least 1 week prior to the experiment and were randomly assigned into different groups.

Administration / exposure

Route of administration:
oral: gavage
CMC (carboxymethyl cellulose)
1% in water
Analytical verification of doses or concentrations:
not specified
Duration of treatment / exposure:
3 days
Frequency of treatment:
once daily
Post exposure period:
Doses / concentrations
Doses / Concentrations:
25 (12,5 M + 12,5 C), 50 (25 M+25C) and 100 (50 M + 50 C) mg/kg bw
actual ingested
No. of animals per sex per dose:
Control animals:
yes, concurrent vehicle
Details on study design:
Thirty minutes before the administration of M+CA, blood samples were collected from the retro-orbital plexus into a vacutainer serum-separating tube (BD Diagnostic, Sparks, MD) for clinical biochemistry using a Selectr E automated analyzer (Vital Scientific, Dieren, Netherlands). For urine collection, rats of the control group and 100 mg/kg/day M+CA-treated group were placed in metabolism cages after the final dose, and urine samples were collected over a 24 h period.
Urine samples were evaluated for creatinine and specific gravity was analyzed with refractometer (ATAGO, Japan). Rats were anesthetized by isoflurane inhalation 24 h after the last dose, and terminal blood samples were obtained from the inferior vena cava for clinical chemistry. Kidneys were removed, weighed, and preserved in 10% neutral-buffered formalin for histopathology.


Histopathology of kidneys
Clinical signs
Changes in body weight
Changes in food consumption
Creatinine clearance
Positive control:

Results and discussion

Details on results:
Visual counting of crystalline deposits shows that rats exposed to a middle dose, 50 mg/kg M+CA, produced renal crystals mainly in the medullar region, while at a high dose, 100 mg/kg M+CA, crystals appeared diffusely throughout the renal cortex and medulla, indicating that M+CA crystals were formed along the osmotic gradient of the kidney. Only the animals with crystals in their kidneys showed increased BUN and creatinine.
Rats given a low dose of 25 mg/kg/day M+CA (12.5 melamine/12.5 cyanuric acid) did not show any evidence of renal impairment or kidney toxicity upon clinical pathologic and histopathologic examination.

Applicant's summary and conclusion