Protein Nitrogen NPN Concentration 224 PHL Lab5 Nonprotein nitrogen NPN NPN includes the nitrogen from all nitrogenous substances other than proteins The NPN could be measured as a ID: 399398
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Slide1
Determination of Serum Non Protein Nitrogen (NPN) Concentration
224 PHL
Lab#5 Slide2
Non-protein nitrogen (NPN)NPN includes the nitrogen from all nitrogenous substances other than proteins.
The NPN could be measured as a
group
or
individually
.
Major Constituents:
Urea, uric acid,
creatinine
, ammonia etc.Slide3
Importance of NPN: Testing
NPN in blood served as a
test for kidney functions
.
Now
it is replaced by determination of
urea nitrogen
because:
(1) The
route of elimination
of various NPN compounds differs considerably.
Some
are excreted by
glomerular filtrations
only e.g.
creatinine
.
Uric
acid
is excreted by
tubular excretion
.
Urea
is excreted by
glomerular filtration
and then partially absorbed by the
tubules
.
(
2) The
increase of NPN
is mainly a reflection of
increase of urea nitrogen
which normally makes up 45% of the total NPN
.Slide4
Determination of Serum Urea ConcentrationUrea
is the end product of
protein metabolism
in the body.
It
is
synthesized in the liver
from the (NH
2
) amino group resulting from
deamination
of amino acids.Slide5
Principle: urease
Urea + H2O ----------> 2NH4 + CO2
Slide6
Procedure:
standard
sample
0.1 ml
Serum sample
0.1 ml
standard
1 ml
1 ml
Working reagent
Incubate for at 37° C for 5 min, then read the absorbance against blank at λ= 520nm.
Normal serum level: 3.5 – 7.2 mg/dl.
* Incubate at
37° C
for
5 min
, then read the absorbance after 30 seconds (A1), then read after 60 seconds (A2) against blank at
λ= 340 nm
.
Normal serum level: 3.5 – 7.2 mg/dl.
,Slide7
Calculation:Serum Urea conc. = A sample
/A
standard
x Conc. Of St. (
53.57)
= mg/dl.
Normal value
: (
10-50
mg/dl)Slide8
Clinical Significance: Increase
in blood urea nitrogen could be due to:
Pre-Renal Causes:
1) Salt and water depletion.
2) Protein catabolism as in fever.
Renal Causes:
1)
Glomerulonephritis
. 2) Mercury poisoning. 3) Hyperparathyroidism. 4)
Hypervitiminosis
D.
cause
↑ in serum Ca and precipitation of Ca in the kidney tissue causing destruction of kidney tissue.Slide9
Clinical Significance: Increase
in blood urea
nitrogen (
hyperuremia
)
could be due to:
Post-Renal
Causes:
1) Prostate enlargement.
2) Stones in urethra. 3) Tumor of the bladder.
cause obstruction to urine flow producing back pressure on the kidney and kidney damage.Slide10
Decrease in blood urea nitrogen (hypouremia) could be due to :
1
) Liver failure.
2
) Malnutrition.
3) Over hydration.
4
) Early stages of pregnancy.Slide11
Determination of Serum Uric Acid Concentration Uric acid is the end product of
purine
metabolism
.
It
comes from endogenous metabolism of nucleoproteins and exogenously from food.Slide12
Principle: uricase
Uric acid + O2 + 2H2O --------->
Alantoin
+ CO2 + H2O2.
peroxidase
2H2O2 + 4-aminoantipyrine + DCFS -------------->
quinoneimine
+ 4H2O.Slide13
Procedure:
standard
sample
0.2 ml
Serum sample
0.2 ml
standard
1 ml
1 ml
Working reagent
Incubate for at 37° C for 5 min, then read the absorbance against blank at λ= 520nm.
Normal serum level: 3.5 – 7.2 mg/dl.
* Incubate at
37° C for 5 min
, then read the absorbance against blank at
λ= 520nm
.
Normal serum level: 3.5 – 7.2 mg/dl.
,Slide14
Calculation:Serum Uric acid conc. = A sample
/A
standard
x Conc. Of St. (6 mg/dl)
= mg/dl.
Normal value
:
3.4
–
7
mg/dlSlide15
Clinical Significance:Increase in uric acid Hyperuricemia
could be due to:
Decrease in uric acid
Hypouricemia
could be due to:
1
.
Gout
.
2. Toxemia.
3. Leukemia.4. Age: menopausal women.5. Drugs:Thiazide diuretics.
1. Hepatitis.
2.
Uricosuric
drugs:
salicylates, phenylbutazone.3. Fanconi
syndrome.Slide16
Determination of Serum Creatinine Concentration :
Creatinine
is the internal anhydride derived from
dephosphorylation
of
creatine
phosphate
.
Creatine
Creatinine + H2O Creatinine has no useful function and is eliminated in urine by glomerular filtration.Slide17
Determination of Serum Creatinine Concentration :
It is
not
reabsorbed by the tubules to any significant extent, Therefore glomerular damage will
decrease the rate at which
creatinine
is excreted.
Creatinine
clearance test
can be used as a
test for kidney function
as its excretion parallels the glomerular filtration rate (GFR).A serum creatinine level over 2 mg/dl indicates
Renal Failure
.Slide18
Principle:Creatinine in the sample reacts with picrate
in alkaline medium forming a colored complex.Slide19
Procedure:
standard
sample
0.1 ml
Serum sample
0.1 ml
standard
1 ml
1 ml
Working reagent
Incubate for at 37° C for 5 min, then read the absorbance against blank at λ= 520nm.
Normal serum level: 3.5 – 7.2 mg/dl.
* Mix then read the absorbance against blank at
λ= 546
nm after 20 seconds (A1).
* Then read after 80 seconds (A2).
Normal serum level: 3.5 – 7.2 mg/dl.
,Slide20
Calculation:Serum Creatinine
conc. = (A2-A1)
sample
/(A2-A1)
standard
x Conc. Of St. (2 mg/dl)
= mg/dl.
Normal value
: (0.6 - 1.1 mg/dl)