Published on February 18, 2014
Amino Acid Disorders Branched-Chain Amino Acid Disorders Purine Disorders Date submitted: February 19, 2014 Section: MD3Y2-7 Name: Pagdanganan, John Kenneth Clanor, Angelica Bonto, Mae Willrose
Amino Acid Disorders ● Amino acid disorders w/ urinary screening tests: ● ● ● ● ● ● ● ● ● Phenylketonuria Tyrosyluria Alkaptonuria Melanonuria Maple syrup urine disease Organic acidemias Indicanuria Cystinuria Cystinosis
Phenylalanine-Tyrosine Disorders ● ● Many of most frequently requested special urinalysis assoc. w/ this metabolic pathway Major inherited disorders: PKU ● Tyrosyluria ● Alkaptonuria Metabolic Defects: Melanoma ● ●
Phenylketonuria ● Most well known aminoaciduria ● 1/10k-20k births ● Can 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
● ● ● 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 retardation ● Alternate pathways develop as child matures ● Aspartame: high levels of phenylalanine
● 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
● 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
● 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
● 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
Tyrosyluria ● Tyrosinemia produces urinary overflow ● Inherited/Metabolic ● Since 2 reactions are directly involved in tyrosin metabolism, urine may have: ● ● Excess tyrosine Excess degraded products: ● P-hydroxyphenylpyruvic acid ● P-hydroxyphenyllactic acid
● 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
● Acquired liver Disease ● Resembles transitory newborn tyrosyluria ● Leucine & Tyrosine crystals seen ● Hereditary Disorders ● ● ● ● ● Absence of enzymes needed in metabolic pathway Serious & usually fatal Liver & Renal Tubular Damage Generalized aminoaciduria 3 types (all produce tyrosylemia & tysosyluria)
● 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
● 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
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
● 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: amber 3. Ehrlich Reagent
Alkaptonuria ● 1 of 6 inborn errors of metabolism ● 3rd major defect in phenylalanine-tyrosine pathway ● ● ● ● ● Absence 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
● 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
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 pathway ● Ketonuria in newborn
Maple Syrup Disease ● Rare, Autosomal recessive trait, caused by IEM ● Involved: leucine, isoleucine, valine ● ● ● Absence 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:
● Newborn screening: MS/MS ● 2,4-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
● 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
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
● Isovaleric Acidemia: ● ● Sweaty feet odor of urine ● Caused by accumulation of isovalerylglycine due to a deficiency of isovaleryl coenzyme A in the leucine pathway Propionic & Methylmalonic acidemia: ● ● Errors converting isoleucine, valine, threonine, methionine to succinyl coenzyme A Immediate precursor to methylmalonic acid: Propionic acid
● Newborn screening: MS/MS ● ● ● ● Isovaleric acidemia Propionic acidemia Methylmalonic acidemia Urine test: Methylmalonic aciduria ● P-nitroaniline: (+) emerald green colour
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
Source: Urinalysis and Body Fluids, 5th edition by Susan King Strasinger & Marjorie Schaub Di Lorenzo
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