ALP

1. RITESH SRIVASTAVA
DEPARTMENT OF BIOCHEMSTRY
SHRIDEV SUMAN SUBHARTI MEDICAL
COLLEGE, DEHRADUN,UTTARAKHAND

2. Phosphatases are enzyme, which catalyze the splitting
of phosphoric acid from certain monophophoric esters,
such as glycerophosphates and phenylphosphates.
There are two type of phosphatase ,i.e.
1. Alkaline phosphatase with a maximum activity at
about pH 10
2. Acid phosphatase, maximum activity between pH 5
and 6.

3. Alkaline phosphatase in serum acts on the substrate
disodium phenylphosphate buffered at pH 10 at 37 ºC
to liberate phenol. This phenol condenses with 4
aminoantipyrine in the presence of alkaline oxidizing
agent potassium ferricyanide to give red coloured
complex which is measured at 520nm.

4. 21-92 IU/L
or
3-13 KA Units
NOTE:- One IU of ALP activity=7.13 KA Units
1KA unit is equal to the production of 1mg phenol by
alkaline phosphatase activity in 100ml of serum in 15
minutes.

5.  Buffered substrate pH-10
 Phenol Std.
 Chromogen Reagent- 4 amino-antipyrine-0.6% &
Potassium ferricyanide-2.4%

6. REAGENT BLANK STD TEST
Buffered Substrate pH-10 0.5ml 0.5ml 0.5ml
D/W 1.55ml 1.55ml 1.55ml
Mix well and Incubate at 37 ºC for 15 min.
REAGENT BLANK STD TEST
Serum ---- ---- 50μl
Phenol STD. ---- 50μl ----
Mix well and Incubate at 37 ºC for 15 min.

7. REAGENT BLANK STD TEST
Colored Reagent (4 aminoantipyrine +
Potassium ferricyanide)
1ml 1ml 1ml
Mix well and read at 520nm.

8. O.D of Blank:-_______________
O.D of STD:-________________
O.D of Test:-_________________
CALCULATION:-
ALP Activity(IU/L) = O.D of Test - O.D of Blank
XConc.
O.D of STD – O.D of Blank Of STD
RESULT:-
INTERPRETATION:-

9. 7/13/2017Biochemistry for Medics 9
 The normal serum alkaline phosphatase consists of many
distinct isoenzymes found in the liver, bone, placenta, and,
less commonly, small intestine.
 Physiological Variations-
 Patients over age 60 can have a mildly elevated alkaline
phosphatase.
 Individuals with blood types O and B can have an elevation of
the serum alkaline phosphatase after eating a fatty meal due to
the influx of intestinal alkaline phosphatase into the blood.
 It is also nonpathologically elevated in children and
adolescents undergoing rapid bone growth, because of bone
alkaline phosphatase.
 It is also high late in normal pregnancy due to the influx of
placental alkaline phosphatase.

10. 1. Hepatic Isoenzyme – Travels fastest towards the
anode .Its level rises in extra hepatic biliary
obstruction.
2. Bone Isoenzyme-Increases due to osteoblastic activity
and is normally elevated in children during periods of
active growth .
3. Placental Isoenzyme - Rises during last 6 weeks of
pregnancy.
4. Intestinal Isoenzyme- Rise occurs after a fatty meal.
May increase during various GI disorders.
7/13/2017Biochemistry for Medics 10

11.  BONE DISEASE:- when osteoblastic activity in bone
formation is take place like in:
 Rickets: values of more than 100KA units can be seen
 Osteomalacia: less rise than in Rickets
 Paget’s Disease: very high values seen
 Healing stage of boon fractures exhibit a transient elevation

12.  LIVER DISEASES:- Enzyme is produced by the
hepatocytes canaliculi and hence marked increase is
seen in condition of obstruction of bile flow.
 Extrahepatic biliary obstruction due to
 stones in bile duct
 Cancer of head of pancreas
 Intrahepatic biliary obstruction due to
viral/durg induced hepatitis or malignancy.
 OTHERS:-
 Fanconi’s syndrome
 Hyperparathyroidism

13.  Severe anemia
 Malnutrition

14. P-nitrophenyl Phosphate + H2O
ALP Mg2+
p-nitrophenol + Phosphate
PROCEDURE
Working Reagent 1000μl
Sample 20μl