NON PROTEIN

NITROGENOUS
COMPOUNDS
KIDNEY FUNCTION TESTS
NON PROTEIN NITROGEN (NPN)
Differ in protein:

CHON high mw
NPN low mw

CHON colloidal
NPN crystals in nature


Renal corpusle : filters blood
Proximal tubule : reabsorbs
electrolytes, nutrients, and
water
Loop of Henle : creates an
osmotic gradient from the
cortex through the medulla
Distal tubule : reabsorbs
elctrolytes and water (under
hormonal control
Collecting duct : gathers all
material that has not returned
to the blood urine
glomerulus Bowmans
capsule
Blood Blood Blood Blood
Filtered
Blood
Filtered
Blood
Filtered
Blood
Filtered
Blood
Blood
solutes
solutes
solutes
solutes
water
water
water
water
water
filtrate
filtrate
filtrate
filtrate
filtrate
filtrate
filtrate
filtrate
filtrate
H
2
O
H
2
O
H
2
O
H
2
O
Na
+
Cl
-
Na
+
Cl
-
Na
+
Cl
-
Urea

Low osmolarity
High osmolarity
Low osmolarity
High osmolarity
urine
FUNCTIONS OF KIDNEYS
EXCRETION
Elimination of metabolic waste products
through the formation of urine
a. glomerular filtration
b. tubular reabsorption
c. tubular secretion
FUNCTIONS OF KIDNEYS
SYNTHETIC
- Erythropoietin
- Renin
- prostaglandins
METABOLIC
- Inactivation of aldosterone, glucagon,
insulin
- Activation of vitamin D
- Formation of creatine
FUNCTIONS OF KIDNEYS
UREA
the major NPN compound in the plasma
Other constituents: (in decreasing order)
(All Underarms Create Cheesy Aroma)
Amino acids
Uric Acids
Creatinine
Creatine
Ammonia
One of the MAJOR PRODUCTS of excretion of
protein catabolism
Formed in the LIVER from CO2 and the ammonia
generated from the deamination of amino acids
Over 90% is excreted, partially reabsorbed along
with water
Readily filtered from the plasma by the glomerulus
UREA
One of the most popular test for assessing renal function
70-80% of glomerular destruction must occur before
there is an increase in the level of plasma urea
Concentration of urea in the plasma is an indicator of
renal function and perfusion, dietary intake of protein
and the level of CHON metabolism
Measurement of plasma urea is further enhanced
when results are considered together with serum
creatinine.

UREA
Clinical significance
AZOTEMIA an elevated concentration of urea in the
blood
Pre-renal
Renal
Post-renal

UREMIA a very high plasma urea concentration
accompanied by renal failure (Uremic syndrome)
PRE-RENAL
related to the renal circulation
the flow of blood to the kidneys
normal renal function
High-protein diet
Muscle wasting ( starvation)
Glucocorticoid treatment
Increased CHON breakdown (stress, fever)
Other related conditions:
RENAL
- involves the kidney there is a lack of ability to function
correctly
- decrease ability to excrete
Polycystic kidneys
http://en.wikipedia.org/wiki/Polycystic_kidney_disease
http://www.futurity.org/tag/polycystic-kidney-disease/
POST-RENAL
Obstruction of the flow of the urine from the kidneys
increase diffusion of urea from the renal tubule into the
circulation

Clinicopathologic Correlation
decreased BUN:
Poor nutrition
High fluid intake
Overhydration (Intravenous fluids)
Pregnancy
Severe liver disease
Analytical Techniques
Specimen: Serum, Plasma, Urine
Specimen not analyzed within few hours should
be refrigerated to prevent bacterial
decomposition
Two (2) analytical approaches:
Direct Method
Indirect Method
UREA = BUN X 2.14
BUN = Urea x 0.467
DIRECT METHOD
DIACETYL MONOXIME Method
- colorimetric
- this method utilizes the Fearon Reaction, wherein urea in hot
solution of diacetyl monoxime condenses to form diazine derivative.
- no interference with ammonia
Urea
+
Diacetyl monoxime
Strong acid
YELLOW
Diazine derivative
Read at 450nm
INDIRECT Method
Berthelot Reaction:
Ammonia is quantitated by spectrophotometric or
electrochemical process
Urea is hydrolyzed to ammonia and bicarbonate ions by urease
ammonia is converted to indophenol with the addition of
nitroprusside
UREA
urease
Ammonia + bicarbonate
Ammonia + nitroprusside indophenol

INDIRECT METHOD
CREATININE
the main storage component of high energy phosphate
needed for muscle metabolism
Synthesize mainly in LIVER from amino acids, arginine,
glycine and methionine
Filtered and secreted, not reabsorbed
Less affected by intake and excretion
Most commonly used in the assessment of GFR
The body content of creatine in normal men is
proportional to muscle mass
CREATININE is formed once creatine losses
water and it is not reused in bodys
metabolism, thus, it is only a waste product
Clinicopathologic Correlation
Increased:
skeletal muscle necrosis or
atrophy.
decrease in glomerular
filtration rate
Creatinine clearance:
CC (mL/min) =
Urine creatinine (mg/dL) x 24 hr urine volume
Serum creatinine (mg/dL) x 1440 (mins in 24 hrs)
Sample Problem
A patient with 24 hour urine
volume is 2000 mL and a
creatinine level of 50 mg/dL. The
serum creatinine is 1.0 mg/dl.
Determine the creatinine
clearance value of the patient.
CC = 50 mg/dl x 2000 ml
1.0 mg/dl x 1440 mins

CC = 69 ml/min
Solution
Analytical Techniques
Specimen: Serum, Plasma, Urine
JAFFE REACTION: most widely used method
Creatinine
+
Alkaline picrate soln
Bright orange-red
complex
(485 nm)
Specimen
Hemolyzed avoided because of
considerable amounts of non-creatinine
chromogens present in RBC
Lipemic and Icteric produce erroneous
results
Maintained at pH 7 during storage
Lloyds reagent remove serum interference
and increases Jaffe Reactions specificity
URIC ACID (urate)
The breakdown product of nucleic
acid and purine catabolism in
humans
At pH 7.4, more than 95% of uric
acid in the body fluids exist as
monosodium urate

UA formation occurs only in tissues that
contain enzyme XANTHINE OXIDASE
Synthesize in the liver and intestinal mucosa
URATE freely filtered by the glomeruli
90% of filtered uric acid is reabsorbed through
active transport
Approximately 8-12% of filtered urate is
excreted in the urine as uric acid
Uric acid in urine exists as:
Monosodium
Disodium
Potassium
Ammonium
Calcium
http://www.udel.edu/mls/mclane/CRCcase2ans4.html
Clinicopathologic Correlation
HYPERURICEMIA serum urate
concentration greater than 7 mg/dl resulting
to numerous disorders and conditions
associated with increased urate production
and/or decreased renal excretion.
GOUT comprises of
heterogeneous group of disorders
Hyperuricemia
Attacks of acute inflammatory
arthritis
Deposition of monosodium urate
crystals through out the body
nephrolithiasis
Specimen: urine, serum
Specimen at room temp: stable for 3 days
Fluoride or thymol: increases stability
UA readily oxidize to allantoin
Chemical and Enzymatic Method:
Caraway Method
Uricase Method
Analytical Techniques
Most methods employ the oxidation of uric
acid
in acid medium allantoin and urea
In alkaline medium allantoin and CO
2

Caraway Method
Based on the oxidation of uric acid by
alkaline phosphotungstate in a CHON-free
solution.
Phosphotungstic acid is reduced to
tungsten blue measured photometrically at
700nm.
Enzymatic Method (uricase)
Oxidation of UA to allantoin using enzyme
uricase (most specific) measured at 293 nm.

UA
uricase
allantoin
AMMONIA
Derived from bacterial action on the
contents of the colon
Metabolize by the liver normally
Increased plasma ammonia is toxic to CNS
Most ammonia is ultimately disposed as
urea
Clinicopathologic Correlation
Altered ammonia metabolism occurs in severe liver
diseases
Elevated in REYEs syndrome
Acute encephalopathy associated with hepatic
dysfunction but without hyperbilirubinemia
Survival reaches to 100% if plasma ammonia
concentration remains below 5 times the normal
AMINO ACIDS
Dietary CHON primary source of AA for
endogenous CHON synthesis
Filtered through the glomerular membrane
Readily reabsorbed in the renal tubules by
active transport
< 5% are excreted in the urine
Clinicopathologic Correlation
Increased urinary excretion of AA fall under two
major types:
OVERFLOW aminoacidurias
Increased urinary excretion with increased plasma
concentration
Acquired secondary or inborn error of metabolism
RENAL aminoacidurias
Diminished tubular reabsorption
Acquired secondary or inborn specific or nonspecific
disorder of the renal tubular reabsorptive mechanism
GUTHRIE Screening Test
Microbial assay
Measures the ability of
phenylalanine in the test sample to
overcome the metabolic inhibition
of -2-thienylalanine on a strain of
Bacillus subtilis

Analytical Techniques
Bacillus subtilis
debivort.org
http://en.wikipedia.org/wiki/File:Phenylketonuria_testing.jpg