1. Amino Acid Disorders Branched-Chain Amino Acid Disorders Purine DisordersDate submitted: February 19, 2014 Section: MD3Y2-7 Name: Pagdanganan, John Kenneth Clanor, Angelica Bonto, Mae Willrose

2. Amino Acid Disorders Amino acid disorders w/ urinary screening tests: Phenylketonuria Tyrosyluria Alkaptonuria Melanonuria Maple syrup urine disease Organic acidemias Indicanuria Cystinuria Cystinosis 3. Phenylalanine-Tyrosine Disorders Many of most frequently requested special urinalysis assoc. w/ this metabolic pathway Major inherited disorders: PKU Tyrosyluria Alkaptonuria Metabolic Defects: Melanoma 4. Phenylketonuria Most well known aminoaciduria1/10k-20k birthsCan cause: Severe mental retardation if undetected Fair complexions due to decreased prod of tyrosine & melanin Identified in Norway by Ivan Folling (1934) Peculiar mousy odour of urine of mentally retarded child High levels of keto acids, inlcuding phenylpyruvate Conversion of phenylalanine to tyrosine is disrupted 5. Absence of gene needed to prod. Phenylalanine hydroxylase Autosomal recessive trait: normal heterozygous carrier Newborn screening test: (+) PKU = eliminate phenylalanine in diet (major constituent in milk) = prevent excessive buildup in serum = avoid mental retardationAlternate pathways develop as child maturesAspartame: high levels of phenylalanine 6. Blood Test Initial screening done b/f being discharged from hospital Can be detected 4hrs after birth if cutoff is lowered to 2mg/dL May need to repeat test in early visit to pediatrician Slower rise in girls- more prone to escape detection during early tests 7. Microbial Inhibition Assay Guthrie Most well-known blood test for PKU P: phenylalanine counteracts action of beta-2thienylalanine, an inhibitor of B. subtilis 1. Blood from heelstick absorbed into filter paper circles 2. Paper disks placed on CM streaked w/ B. Subtilis (+) growth around paper disk (-) no growth 8. Urine Test Appears after 2-6 weeks in urine Follow up procedure in questionable diagnostic cases Screening test to ensure proper dietary control Monitoring dietary intake of pregnant women lacking phenylalanine hydroxylase 9. Ferric Chloride Urine tube test testing for phenylpyrvic acid Nonspecific: reacts with other amino acids & medications False (+) produced by certain brands of disposable diapers Permanent blue-green colour 1. Place 1mL of urine in tube 2. Slowly add 5 drops of 10% ferric chloride 3. Observe colour 10. Tyrosyluria Tyrosinemia produces urinary overflowInherited/MetabolicSince 2 reactions are directly involved in tyrosin metabolism, urine may have: Excess tyrosine Excess degraded products: P-hydroxyphenylpyruvic acid P-hydroxyphenyllactic acid 11. Transitory Tyrosinemia Most frequently seen in premature infants Caused by underdevelopment of liver fxn req. to prod. enzymes needed for complete metabolism Rarely causes permanent damage Difference in urinary test w/ PKU Green colour fades away quickly 12. Acquired liver Disease Resembles transitory newborn tyrosyluriaLeucine & Tyrosine crystals seenHereditary Disorders Absence of enzymes needed in metabolic pathway Serious & usually fatal Liver & Renal Tubular Damage Generalized aminoaciduria 3 types (all produce tyrosylemia & tysosyluria) 13. Type 1: Deficient in Fumarylacetoacetate hydrolase (FAH) Generalized renal tubular disorder & progressive liver failure Type 2: Deficient in Tyrosine aminotransferase Corneal erosion; lesions of palms, fingers, & soles of feet due to crystallization of tyrosine in cells Type 3: Deficient in P-hydroxyphenylpyruvic acid dioxygenase Mental retardation if dietary restrions of phenylalanine & tyrosine not implemented 14. Nitroso-naphtol Test Urinary screening test for tyrosine & its metabolites Nonspecific (+) orange-red = indicates need of further tests 1. Place 5 drops of urine in a tube 2. Add 1mL of 2.63N nitric acid 3. Add 1 drop of 21.5% sodium nitrite 4. Add 0.1mL l-nitroso-2-napthol 5.Mix 6. Wait 5 minutes 7. Observe colour MS/MS screening test Available for Type 1 & 2 15. Melanuria 2nd pathway of Tyrosine Responsible for prod. of melanin, thyroxine, epinephrine, protein & tyrosine sulfate Major lab concern: Melanin Confers dark hair, eye, and skin colour Deficieny: Albinism Elevated: overproliferation of melanocytes = Malignant Melanoma Tumor cells secrete 5,6-dihydroxyindole = oxidizes to melanogen, then melanin = dark urine 16. Melanin reacts w/: 1. Ferric Chloride: gray/black precipitate 2. Sodium Nitroprusside (nitroferricyanide): red Glacial acetic acid: prevents intereference from creatinine & acetone Melanin: green-black Acetone: purple Creatinine: amber3. Ehrlich Reagent 17. Alkaptonuria 1 of 6 inborn errors of metabolism3rd major defect in phenylalanine-tyrosine pathwayAbsence of gene needed to prod. Homogentisic acid oxidase Homogentisic acid accumulates in blood, tissue, urine alkali lover - turns dark & alkali after standing in room temp Brown pigment deposited in body tissues (esp. ear); deposit in catilage (arthritis) Many develop liver & cardiac disorders 18. Homogentisic acid reacts w/: 1. Ferric Chloride: transient deep blue colour 2. Clinitest: yellow precipitate 3. Alkali: darkening of urine (interference: large amounts of ascorbic acid) Silver nitrate & ammonium hydroxide: black Read: Spectrophotometer, Chromatography procedures 19. Branched-Chain Amino Acid Disorders A methyl group branches from main aliphatic carbon chain 1. accumulation of 1/more early amino acid degradation products (maple syrup urine dse) 2. organic acidemias: accumulation of organic acids further down metabolic pathwayKetonuria in newborn 20. Maple Syrup Disease Rare, Autosomal recessive trait, caused by IEMInvolved: leucine, isoleucine, valineAbsence of gene for oxidative carboxylation of keto acids (alpha-ketoisovaleric, alpha-ketoisocaproic, alpha-keto-beta-methylvaleric) = accumulation in blood & urine Failure to thrive after aprox. 1 week Maple syrup odor (11th day): due to rapid accumulation of keto acids REPORT! Prevent severe mental retardation & death: 21. Newborn screening: MS/MS2,4-dinitrophenylhydrazine (DNPH) reaction Most frequently performed urinary screening (+) yellow turbidity or precipitate Interferece: Large doses of ampicillin Nonspecific Treatment can be started based on odor, clinical symptoms & positive DNPH test while confirmatory prodecures are being done 22. 2,4-Dinitrophenylhydrazine Test 1. Place 1mL of urine in a tube 2. Add 10 drops of 0.2% 2,4-DNPH in 2N Hcl 3. Wait 10 minutes 4. Observe for yellow/white precipitate 23. Organic Acidemias Generalized symptoms: Early severe illness Vomiting w/ Metabolic acidosis Hypoglycemia Ketonuria Increased serum ammonia Most frequent disorders: Isovaleric acidemia Propionic acidemia Methylmalonic acidemia 24. Isovaleric Acidemia: Sweaty feet odor of urine Caused by accumulation of isovalerylglycine due to a deficiency of isovaleryl coenzyme A in the leucine pathwayPropionic & Methylmalonic acidemia: Errors converting isoleucine, valine, threonine, methionine to succinyl coenzyme A Immediate precursor to methylmalonic acid: Propionic acid 25. Newborn screening: MS/MS Isovaleric acidemia Propionic acidemia Methylmalonic acidemiaUrine test: Methylmalonic aciduria P-nitroaniline: (+) emerald green colour 26. Purine Disorders Lesch-Nyhan Disease Sex linked recessive Massive excretion of uric acid crystals Absence of Hypoxanthine guanine phosphoribosyltransferase Severe motor defects, mental retardation, tendency towards self-destruction, gout, renal calculi First 6-8 months: normal development First symptom: (uric acid) orange sand in diapers 27. Source: Urinalysis and Body Fluids, 5th edition by Susan King Strasinger & Marjorie Schaub Di Lorenzo