NPN Flashcards
UREA
Enzymatic Method
Chemical Method
GLDH coupled enzyme
Indicator dye
Conductimetric (ISE)
Fearon’s reaction
UREA
First step
Urea + 2 H2O –Urease 2 NH4+ + CO3 2-
GLDH coupled enzyme disappearance of NADH at ___
NH4+ + 2-oxoglutarate + NADH + H+ _____ glutamate + NAD+ + H2O
340 nm
NH4+ + 2-oxoglutarate + NADH + H+ GLDH glutamate + NAD+ + H2O
Indicator dye
NH4+ + pH Indicator color change
Nessler’s reaction (HgI2/KI)
Berthelot’s reaction (Alk. hypochlorite)
Conductimetric (ISE)
Conversion of unionized urea to NH4+ and CO32- results in increased conductivity
Fearon’s reaction
Urea + DAM (____.) ____ (Diazine dirivative)
Urea + DAM (Diacetyl Monoxime Mtd.) Yellow solution (Diazine dirivative)
Plasma or serum
Urine, 24 hour
6 – 20 mg/dL - 2.1-7.1 mmol/L
12-20 g/day - 11.5-4.4 mmol urea/day
Specimen Considerations
- Use fasting blood
- Avoid fluoride or citrate anticoagulants
- Refrigerate Samples
(↑ urea in the blood)
Azotemia
Azotemia 3
Prerenal
Renal (Uremia)
Postrenal
Reduce blood flow or ↑ protein catabolism
Prerenal
Damage of nephrons (Renal failure/disease)
Renal (Uremia)
Urinary tract obstruction (calculi, tumors)
Postrenal
Azotemia
4 Decreased Concentration
Low protein intake
Severe vomiting and diarrhea
Liver disease
Pregnancy
URIC ACID
1. Physiology
Major end-product of purine catabolism.
Present as monosodium urates in plasma (98-100% reabsorbed)
May precipitate in tissues (blood pH ≈ 7 ; BUA > 6.8 mg/dL )
URIC ACID
2. Clinical Application
Inherited disorders of purine metabolism Gout Renal calculi Uric acid nephropathy (chemotheraphy) Kidney dysfunction
URIC ACID
Enzymatic Methods
Chemical Method
Spectrophotometric
(Blauch and Koch)
Coupled enzyme (I) Catalase
Coupled enzyme (II) Peroxidase
Phosphotungstic acid
(Caraway method)
URIC ACID
First step
Uric Acid + O2 +2 H2O –____ ____ + CO2 + ____
Uric Acid + O2 +2 H2O –Uricase allantoin + CO2 + H2O2
Spectrophotometric
(Blauch and Koch)
↓ absorbance at ___
↓ absorbance at 293 nm
Uric acid v. allantoin
Coupled enzyme (I)
Catalase
H2O2 + reagent –____ colored compound
H2O2 + reagent –Catalase colored compound
Coupled enzyme (II) Peroxidase H2O2 + indicator dye –\_\_\_\_\_ - \_\_\_\_
H2O2 + indicator dye –Peroxidase Red dye
Phosphotungstic acid
(Caraway method)
Uric Acid + H3PW12O4o + O2
- ______ allantoin + ____ + CO2
Uric Acid + H3PW12O4o + O2
- Na2CO3/OH-
allantoin + tungsten blue + CO2
Specimen Considerations
urea 3
- Heparinized plasma, serum or urine
- Lipemia, bilirubin, hemolysis (↓ UA)
- Salicylates and thiazides (↑ UA)
Hyperuricemia (Enzyme deficiencies)
1) Lesch-Nyhan syndrome
2) Glycogen storage disease type 1
3) Fructose intolerance
4) Hemolytic and proliferative process
5) Treatment of myeloproliferative disease w/ cytotoxic drugs
6) Chronic renal disease
7) Toxemia of pregnancy and lactic acidosis
8) Purine-rich diet
Lesch-Nyhan syndrome
↓ HGPRT
↓ reutilization of purine, ↑ synthesis of purine
Glycogen storage disease type 1
↓ G-6-P
↑ triglycerides, ↓ renal urate excretion
Fructose intolerance
↓ F1PA
↑ lactate, ↓ renal urate excretion
Hypouricemia
5
↓ PRPP Synthetase Liver disease Fanconi sydrome Chemotheraphy with azathioprine or 6-mercaptopurine Overtreatment with allopurinol
↓ PRPP Synthetase
↓ de novo purine synthesis
Liver disease
↓ uric acid synthesis
Fanconi sydrome
Defective tubular reabsorption
Chemotheraphy with azathioprine or 6-mercaptopurine
↓ de novo purine synthesis
Overtreatment with allopurinol
↓ XO, ↓ de novo purine synthesis
CREATININE
Physiology
Chief product of muscle metabolism
Not affected by protein diet
CREATININE
2. Clinical Application
Sufficiency of kidney function
Severity of kidney damage
Progression of kidney disease
Completeness of 24-hour urine
Volume of plasma filtered (V) by the glomeruli per unit of time (mL/minute)
Glomerular filtration rate (Creatinine clearance)
GFR = V/t
GFR = UCrVu
———— x 1.73/A
PCrt
CREATININE
Chemical Method
Jaffe Reaction
Jaffe-kinetic
Jaffe with adsorbent
CREATININE
Principle
Creatinine + picrate Red-orange complex
CREATININE
Jaffe-kinetic
Rate of change of absorbance(color formation)
Principle
Jaffe with adsorbent
PFF is adsorbed first onto Fuller’s earth (aluminum magnesium silicate)
CREATININE
6
Falsely ↑
Glucose α-ketoacids Ascorbate Uric Acid Cephalosporins Dopamine
CREATININE
3
Falsely ↓
- Bilirubin
- Hemoglobin
- Lipemic specimens
CREATININE
Increased Concentration
Renal failure (glomerular function) ↑ Plasma Concentration = ↓ GFR
AMMONIA
1. Physiology
By product of amino acid deamination.
Converted to urea in the liver.
AMMONIA
2. Clinical Application
Hepatic failure and hepatic coma
Reye’s syndrome
Inherited deficiencies of urea cycle
AMMONIA
Chemical Method
Enzymatic Method
Ion-selective electrode
Spectrophotometric
GLDH
↓ absorbance (340 nm)
Chemical Method
Principle
-Ion-selective electrode
Diffusion of ___ through selective membrane into ___ causing pH change.
-Spectrophotometric
-Enzymatic Method
GLDH
NH4+ + 2-oxoglutarate + ____ + H+ ____ Glutamate + NADP+ + H2O
NH3
NH4Cl
NH3 + bromphenol blue blue dye
NH4+ + 2-oxoglutarate + NADPH + H+ GLDH Glutamate + NADP+ + H2O
AMMONIA
Specimen Considerations
- Heparinized and EDTA tubes
- Samples is centrifuged at 0-4°C within 20 mins. of collection and the plasma/serum removed
- Avoid cigarette smoking for several hours
