Quantification of N-acetylaspartic acid in urine by LC-MS/MS for the diagnosis of Canavan disease.

[canavan disease]

Canavan disease is an autosomal recessive leukodystrophy characterized by excessive excretion of N-acetylaspartic acid (NAA ) in urine . The disease is caused by deficiency of aspartoacylase , the enzyme responsible for the hydrolysis of NAA into acetate and l-aspartate . Patients , who are often asymptomatic in their early months , show a wide spectrum of clinical presentation thereafter that includes macrocephaly , poor head control , seizures , abnormal muscle tone , optic atrophy , significant developmental delay and death . In this work , we describe a simple liquid chromatography-tandem mass spectrometry (LC -MS /MS ) method for the determination of NAA in urine . The internal standard d 3 -NAA was added to untreated urine and the mixture was injected into the LC -MS /MS system operated in the negative ion mode . Detection was achieved in multiple reaction monitoring (MRM ) mode by monitoring m /z 174 --> 88 , 174 --> 130 and 174 --> 58 for NAA and 177 --> 89 for the internal standard . Separation was carried out on a C 8 column (2 .1 x 150 mm ) using a mixture of acetonitrile and water (1 :1 v /v ) containing 0 .05 % formic acid at a flow rate of 0 .25 ml /min . NAA was eluted at 1 .6 min and the run time was approximately 2 min . Using spiked urine , the assay was linear up to 2 mmol /L with limit of quantification at 1 micromol /L (S /N = 12 ). NAA in patients ' urine (n = 17 ) ranged between 366 and 21 ,235 mmol /mol creatinine compared to controls of <39 mmol /mol creatinine (n = 159 ). This LC -MS /MS method for NAA as described involved no extraction and no derivatization , showed no interference , and gave excellent recovery with low variability and short analytical time .

Diseases
Validation

Diseases presenting "deficiency of aspartoacylase" symptom

canavan disease

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