Melamine Toxicity

Melamine Melamine, (2,4,6-triamino- s -triazine), is an industrial chemical best known for its use in brightly colored dinnerware plastics since the 1950s coined with the name M e lmac dishes. It is also used as a fertilizer in Asia and often added to wheat flour because of its high nitrogen content to increase the apparent protein content. Melamine has been extensively evaluated for toxicity; it does not undergo significant metabolism and for the most part is readily cleared via the kidney within 24 hours of consumption.

Melamine, (2,4,6-triamino- s -triazine), is an industrial chemical best known for its use in brightly colored dinnerware plastics since the 1950s coined with the name M e lmac dishes.

It is also used as a fertilizer in Asia and often added to wheat flour because of its high nitrogen content to increase the apparent protein content.

Melamine has been extensively evaluated for toxicity; it does not undergo significant metabolism and for the most part is readily cleared via the kidney within 24 hours of consumption.

Recent News about Melamine Tainted milk and milk products in China in September and October 2008 54,000 children sickened and at least 4 fatalities Resonates with the melamine poisoning of cat and dog food in March 2007

Tainted milk and milk products in China in September and October 2008

54,000 children sickened and at least 4 fatalities

Resonates with the melamine poisoning of cat and dog food in March 2007

Cats and dogs were dying from acute renal failure. Melamine and other melamine-related triazines were found in the wheat gluten that Menu Foods® was using from China. Melamine alone is essentially nontoxic with an LD 50 in rats of 3161 mg/kg. Much less is known about the other melamine-related compounds and their toxicity. A month after the melamine was discovered, the FDA announced that cyanuric acid was also found in the pet food which also has a high LD 50 in rats of 7700 mg/kg.

Cats and dogs were dying from acute renal failure.

Melamine and other melamine-related triazines were found in the wheat gluten that Menu Foods® was using from China.

Melamine alone is essentially nontoxic with an LD 50 in rats of 3161 mg/kg. Much less is known about the other melamine-related compounds and their toxicity.

A month after the melamine was discovered, the FDA announced that cyanuric acid was also found in the pet food which also has a high LD 50 in rats of 7700 mg/kg.

Researchers had to figure out then what caused the acute renal failure. Was it one of the other melamine-related compounds that was causing the toxicity? Was it some combination of the melamine, cyanuric acid, or other triazines?

Researchers had to figure out then what caused the acute renal failure.

Was it one of the other melamine-related compounds that was causing the toxicity?

Was it some combination of the melamine, cyanuric acid, or other triazines?

Identification and Characterization of Toxicity of Contaminants in Pet Food Leading to an Outbreak of Renal Toxicity in Cats and Dogs Roy L. M. Dobson, Safa Motlagh, Mike Quijano, R. Thomas Cambron, Timothy R. Baker, Aletha M. Pullen, Brian T. Regg, Adrienne S. Bigalow-Kern, Thomas Vennard, Andrew Fix, Renate Reimschuesse, Gary Overmann, Yuching Shan and George P. Daston Proctor and Gamble Toxicological Sciences 2008 106(1):251-262

Roy L. M. Dobson, Safa Motlagh, Mike Quijano, R. Thomas Cambron, Timothy R. Baker, Aletha M. Pullen, Brian T. Regg, Adrienne S. Bigalow-Kern, Thomas Vennard, Andrew Fix, Renate Reimschuesse, Gary Overmann, Yuching Shan and George P. Daston

Proctor and Gamble

Toxicological Sciences 2008 106(1):251-262

Cytotoxicity Testing Madin-Darby canine kidney cells and Crandell feline kidney cells in vitro study of each triazine None of the chemicals alone caused any significant cytotoxicity

Madin-Darby canine kidney cells and Crandell feline kidney cells in vitro study of each triazine

None of the chemicals alone caused any significant cytotoxicity

Copyright restrictions may apply. Dobson, R. L. M. et al. Toxicol. Sci. 2008 106:251-262; doi:10.1093/toxsci/kfn160 Cytotoxicity data for six triazines and a positive control (mercuric chloride) in Madin-Darby canine kidney cells (diamonds) and Crandell feline kidney cells (squares)

Acute In Vivo Study Ammeline and ammelide were given by gavage to female Sprague-Dawley rats in 10, 30, and 100 mg/kg doses. Clinical chemistry and histopathology of blood, urine, and tissues samples showed no signs of toxicity thus ruling out that one of the melamine-related compounds was sufficiently toxic enough to cause the acute renal failure.

Ammeline and ammelide were given by gavage to female Sprague-Dawley rats in 10, 30, and 100 mg/kg doses.

Clinical chemistry and histopathology of blood, urine, and tissues samples showed no signs of toxicity thus ruling out that one of the melamine-related compounds was sufficiently toxic enough to cause the acute renal failure.

2nd Acute In Vivo Study Two different groups of female Sprague-Dawley rats were treated with a combination of either melamine, ammeline, and ammelide or with melamine and cyanuric acid. Initial diuresis resulted followed by oliguria especially in the melamine-cyanuric acid group. Clinical chemistry of blood samples showed an increase in blood urea nitrogen levels and a decrease in creatinine clearance, both of which are indicators of renal impairment.

Two different groups of female Sprague-Dawley rats were treated with a combination of either melamine, ammeline, and ammelide or with melamine and cyanuric acid.

Initial diuresis resulted followed by oliguria especially in the melamine-cyanuric acid group.

Clinical chemistry of blood samples showed an increase in blood urea nitrogen levels and a decrease in creatinine clearance, both of which are indicators of renal impairment.

Copyright restrictions may apply. Dobson, R. L. M. et al. Toxicol. Sci. 2008 106:251-262; doi:10.1093/toxsci/kfn160 Urine volume (measured on a daily basis) in rats receiving either a mixture of melamine/ammeline/ammelide/cyanuric acid (400/40/40/40 mg/kg/day), or melamine/cyanuric acid (400/400 mg/kg/day) for three days

2nd In Vivo Study (Cont’d.) Histological examination of the kidney tubules revealed brownish-yellow crystals sufficiently large and abundant enough to block tubular flow, with little cellular damage, consistent with the kidneys of donated pets exposed to this food.

Histological examination of the kidney tubules revealed brownish-yellow crystals sufficiently large and abundant enough to block tubular flow, with little cellular damage, consistent with the kidneys of donated pets exposed to this food.

Copyright restrictions may apply. Dobson, R. L. M. et al. Toxicol. Sci. 2008 106:251-262; doi:10.1093/toxsci/kfn160 (a) Gross morphology of rat kidneys after three days of treatment with a mixture of four triazines (mixture) or melamine and cyanuric acid

Copyright restrictions may apply. Dobson, R. L. M. et al. Toxicol. Sci. 2008 106:251-262; doi:10.1093/toxsci/kfn160 Kidney histology in the cat that had died after eating contaminated food (a, c) and a rat treated with melamine/cyanuric acid (b, d)

2nd In Vivo Study (cont’d.) Crystals from the rat kidneys and from donated cat kidneys from veterinarians, in addition to crystals isolated from the wheat gluten directly, were evaluated using FTIR spectroscopy. The spectra collected from these samples matched the reference spectrum of a melamine-cyanuric acid reference material.

Crystals from the rat kidneys and from donated cat kidneys from veterinarians, in addition to crystals isolated from the wheat gluten directly, were evaluated using FTIR spectroscopy.

The spectra collected from these samples matched the reference spectrum of a melamine-cyanuric acid reference material.

Copyright restrictions may apply. Dobson, R. L. M. et al. Toxicol. Sci. 2008 106:251-262; doi:10.1093/toxsci/kfn160 Comparison of FTIR spectra collected from a particle isolated from the suspect wheat gluten (top), a crystal located within the cat kidney (middle), and a melamine-cyanuric acid cocrystal reference compound (bottom)

Conclusion Because the in vitro study of the individual chemicals caused no significant cytotoxicity, and because the kidneys themselves showed no cellular damage, it is concluded that the acute renal toxicity is due to physical blockage by melamine-cyanuric acid crystals.

Because the in vitro study of the individual chemicals caused no significant cytotoxicity, and because the kidneys themselves showed no cellular damage, it is concluded that the acute renal toxicity is due to physical blockage by melamine-cyanuric acid crystals.