NON-PROTEIN NITROGENS

LARAMELISAR.OLGUERA, RMT, MSMT©, MT (AMT), MLS(ASCPi)CM

BASICand FUNDAMENTALS
Urine Formation
• Ultrafiltrate of plasma.
• Conversion of 170,000 mL of
filtered plasma to 1,200 mL of
urine.
• Kidneys continuously form urine
through:
✓Filtration
✓Reabsorption
✓Secretion
BASICand FUNDAMENTALS
• Urine Composition
• General composition:
✓Urea
✓Organic & inorganic chemicals
✓95%water; 5% solute
BASICand FUNDAMENTALS
• Urine Composition
ORGANIC (35 grams) INORGANIC (25 OTHERS
grams)
Urea: PRIMARY Chloride: PRIMARY Hormones,
Vitamins,
Creatinine Sodium Medications, Formed
elements (cells,
Uric Acid Potassium casts, crystals, etc.)
UREA
• Major end product of protein (dietary) catabolism; 45% of total NPN.
• Synthesis: Liver from CO2 and the ammonia from the deamination.
• By-product of Ornithine or Kreb’s Henseleit Cycle
• about 25 grams of Urea is excreted daily
• concentration of Urea BUN: 2.14 x BUN= urea (mg%)
• BUN:Creatinine Ratio =10:1 – 20:1
• NV: 8-23 mg/dL
CREATININE
• End product of MUSCLE METABOLISM derived from CREATINE
• Chemical name of Creatine: α-methyl guanidoacetic acid
• Produced by 3 amino acids: Methionine, Arginine, Lysine
• Index of overall renal function
• Amount generated =SKELETAL MUSCLE PRESENT
• NV: M: 0.9-1.3 mg/dL, F: 0.6-1.1 mg/dL
ACUTEKIDNEYINJURYCLASSIFICATION
STAGE SERUM CREATININE CRITERIA URINE OUTPUT CRITERIA

1 >0.3 mg/dL or 150%-200% <0.5 mL/kg for >6hr

2 >200% - 300% <0.5 mL/kg for>12hr

3 >4mg/dL, 300%, or acute <0.3 mL/kg for >24

increase of >0.5mg/dL hr or anuria >12hr
SERUM URIC ACID
• Major product of PURINE CATABOLISM
• Formation: from xanthine by the action of xanthine oxidase in the liver
and intestine.
• Weak acid; pH 7.4; 95% or more are Monosodium Urate
• About 1 gram is excreted daily
• NV: Uricase M: 3.5 – 7.2 mg/dL F: 2.6 – 6.0 mg/dL
INCREASED SERUM URIC ACID
• HYPERURICEMIA • HYPORURICEMIA
• GOUT • Fanconi’s syndrome
• INCREASED NUCLEAR • Wilson’s disease
METABOLISM • Hodgkin’s disease
• Chronic kidney disease
• Lesch-Nyhan syndrome
AMMONIA
• Product of PROTEIN
DEAMINATION
• increases in HEPATIC COMAand
REYE’S SYNDROME
• Toxic substance: converted to
urea by the liver then excreted
into the kidney.
• SPECIMEN CONSIDERATION:
• Eliminate source of contamination
• EDTA Plasma or Heparinized
Plasma; SERUM IS NOT USED!
• Place on Ice Bath and Analyze
Immediately!
• Considered as STAT SPECIMEN
METHODS: BUN
1. Chemical Method (Direct) • SPECIMEN CONSIDERATION:
a.Diacetyl Monoxime Method (Fearon • Fasting NOT REQUIRED
Reaction)
• Fluoride or Citrate inhibit urease
UREA+ DAM → Yellow Diazine
Derivative • Color developer:
Thiosemicarbazide and Ferric
b. O-phthaldehyde Ions
UREA +o-phtaldehyde →
ISOINDOLINE + ETHYLENE DIAMINE
→ Colored production
(+) measured spectrophotometrically
METHODS: BUN
1. Enzymatic Method (Indirect)
a. Urease Method
• urease prepared from JACK BEANS
• after urease; ammonia can be treated with BERTHELOT
• Ammonia and CO2 are measured; NH3 most often

b. Couple Urease/Glutamate Dehydrogenase Method - UV Enzymatic

Primary Reaction:
METHODS: BUN
2. ENZYMATIC METHOD
b. Couple Urease/Glutamate Dehydrogenase Method- UV Enzymatic
Secondary Reaction:
METHODS: BUN
2. ENZYMATIC METHOD
b. Couple Urease/Glutamate Dehydrogenase Method- UV Enzymatic
Secondary Reaction:
METHODS: CREATININE
SPECIMEN CONSIDERATION:
• Fasting NOT REQUIRED
• Avoid HEMOLYZED and ICTERICSAMPLES
• Index of Renal Function: Serum and Urine Creatinine
• Cephalosporin: FALSEINCREASE IN JAFFE REACTION

• TAKE NOTE! 24-Hour Urine Sample with <0.8 g/day of creatinine

indicates that some of the urine was probably discarded.
METHODS: CREATININE
Chemical Method – Direct Jaffe Method
• Creatinine +Alk. Picrate → RED-ORANGE TAUTOMER of Creatinine
• Interference:

FALSE INCREASED FALSE DECREASED

Ascorbic Acid, Glucose, Bilirubin and hemoglobin
Uric Acid and α-keto acids
METHODS: CREATININE
• Folin Wu Method - sensitive; NON-SPECIFIC
• Lloyd or Fuller’s Earth Method - sensitive; SPECIFIC [MODIFIED
JAFFE].
• Adsorbents:
1. Lloyd’s Reagent- Sodium Aluminum Silicate
2. Fuller’s Earth Reagent- Aluminum Mg2+ Silicate
• JAFFE REAGENT: ALKALINE PICRATE
✓Saturated Picric Acid
✓10%NaOH
METHODS: CREATININE
• Kinetic Jaffe Reaction
✓requires PRECISION EQUIPMENT; rapid and easy
• Reaction:
• Alk. Picrate +Serum (Crea) → measure absorbance
• Janovsky-Like Reaction- Carbonyl oxygen of creatinine may attack the
I-carbon of Picric forming covalent adduct
• Enzymatic Reaction
• Isotope Dilution Mass Spectrometry (IDMS) – REFERENCEMETHOD
METHODS: URICACID
• Chemical Methods: Oxidation-Reduction Reaction

a. Sodium Cyanine (NaCN) =Folin, Brown, Newton Benedict

b. Sodium Carbonate (Na2CO3) =Archibald, Henry, Caraway

• Enzymatic Methods
• URICASE METHOD: (SUA has UV abs. peak at 293nm, allantoin doesn’t
have)
METHODS: URICACID
• Isotope Dilution Mass Spectrometry IDMS) – REFERENCE METHOD
• SPECIMEN CONSIDERATION:
✓Fasting not required but preferred
✓Stable in urine and serum for 3 days; Room Temp
✓Do not use Potassium Oxalate
✓Interferences: Ascorbic Acid and Bilirubin
DISEASEASSOCIATION
GENERAL SPECIFIC

Azotemia – Pre-Renal – KIDNEY IS NOT THE PROBLEM; Overflow of Analyte in the BLOOD
✓ Decreased Renal Blood Flow; BUN ↑;Plasma Creatinine Normal
NITROGENO
US Renal – Kidney problem; inability to convert NPN into eliminable substance
SUBSTANCE ✓ Striking BUN Level; Slow Rising of Plasma Creatinine, Anemia, Electrolyte Balance
increase Post Renal – Urinary tract obstruction (Low GFR); Renal Calculi, Cancer
✓ Urea higher than Creatinine (back diffusion of urea in circulation)
DISEASEASSOCIATION
GENERAL SPECIFIC

Uremia • severe form of azotemia; basic azotemia symptoms + acidemia and K+ Elevation
• Normo-Normo, Uremic Frost (dirty Skin), Edema, FOUL BREATH AND URINE-LIKE
SWEAT
Hyperuricemia a. Gout- definitive diagnosis: “birefringent crystal” in synovial fluid
b. Allapurinol drug
c. Chronic Renal Disease: BUA: >10mg/Dl (RENAL CALCULI)
d. Lesch-Nyhan Syndrome: HGPRT deficiency