CC2 AMYLASE

• AMYLASE

  Also known as:

  1.Diastase

  2.Alpha 1,4-glucan-4-gluconhydrolase 3. 1,4alpha-D Glucanohydrolase. 4. EC: 3.2.1.1
• An enzyme belonging to the class of hydrolases that catalyze the breakdown of starch and glycogen (complex carbohydrate) into sugars
• Low molecular weight enzyme that can be cleared in the urine
• • Action:
  • catalyzes the breakdown of starch and glycogen via hydrolysis to form maltose and maltodextrin
  • Maltose -being the major end product
• • a-amylase attacks only at the a, 1-4 glycosidic bond to produce degradation products consisting of: a. glucose b.maltose- major end product

  c. intermediate chains called dextrines which contain alpha, 1-6 branching linkages
• Starch consists of both amylose and amylopectin
• AMYLOSE

  -is long

  -unbranched chain of glucose molecules

  • linked by a, 1-4 glycosidic bonds
  • soluble in water
• AMYLOPECTIN

  • a branched-chain polysaccharide
  • with a, 1-6 linkages at the branched points
  • insoluble fraction
• structure of glycogen is similar to that of amylopectin but is more highly branched
• Amylase

  • An important enzyme in the physiologic digestion of starches
• Amylase

  • Requires activators for its activity, namely: calcium and chloride
• Types of Amylase:

  Alpha Amylase (EC.3.2.1.1)

  Beta Amylase (EC.3.2.1.2)

  Gamma Amylase (EC3.2.1.3)
• Alpha Amylase (EC 3.2.1.1)

  • 1,4-a-D-glucan glucanohydrolase/ glycogenase
  • Calcium metalloenzymes
  • Found in animal tissues and fluids
  • Most stable enzyme that is of clinical interest
• Beta Amylase (EC 3.2.1,2)1,4-a-D-glucan maltohydrolase/glycogenase/saccharogen amylase. Found in both plants and animals. Acts on fragments of starch molecule
• Gamma Amylase (EC 3.2.1 3)

  Glucan 1,4-glucosidase/ amyloglucosidase/ Exo-1,4-a-glucosidase/glucoamylase/ lysosomal a-glucosidase/ 1,4-a-D-

  glucangluhydrolase

  • Found in numerous fungi
  • Will cleave a (1-6) glycosidic linkages and a (1-4) glycosidic linkages at the nonreducing end of amylose and amylopectin, yielding glucose. Has most acidic optimum pof all amylases because it is more active around рн3
• Gamma Amylase (3.2.1.3)

  • Has most acidic optimum pof all amylases because it is more active around рн3
• • 2 Major Sources of the AMS. 1.) Acinar cells of the pancreas 2.) Salivary glands-parotid gland
• • The smallest enzyme (MW: 50,000-55,000 Daltons)
• • Digestion of starches begins in the mouth with the hydrolytic action of salivary amylase (ptyalin). Salivary activity, however, is of short duration because upon swallowing it is activated by the acidity of the gastric contents.
• • Digestion of starches begins in the mouth with the hydrolytic action of salivary amylase (ptyalin). Salivary activity, however, is of short duration because upon swallowing it is activated by the acidity of the gastric contents. Pancreatic amylase then perform the major digestive action of starches once the polysaccharide reaches the intestine.
• Two (2) types of Isoenzymes

  1. P type Isoamylase (P1, P2, and P3)

  • From pancreatic tissue
  • Slow moving in CHO electrophoresis
  • Predominates in normal urine

  2. S type Isoamylase (S1, S2, and S3)

  • From salivary gland tissue as well as fallopian tube and lung. • Fast moving/migrate most quickly
  • Approximately 2/3 of AMS activity of normal serum
• P type Isoamylase (P1, P2, and P3)

  • From pancreatic tissue
  • Slow moving in CHO electrophoresis. > Predominates in normal urine
• S type Isoamylase (S1, S2, and S3)

  • From salivary gland tissue as well as fallopian tube and lung
  • Fast moving/migrate most quickly
  • Approximately 2/3 of AMS activity of normal serum
• • P1 Isoenzyme
  • rarely observed fraction
• P2, S1, and S2

  • most observed fractions
• S1

  -slowest migrating salivary band
• P2

  • prominent peak
  • intermediate migrating pancreatic bond
• P3

  • prominent in acute pancreatitis
  • frequently occurs in patients w/ severe kidney diseases and not entirely specific
• • Quantitation is done through: 1. Electrophoresis- uses cellulose acetate 2. Column chromatography - provides most reliable data in research purposes 3. Isoelectric focusing 4. Wheat Germ Inhibition Method- using Wheat Germ Inhibitor by Beckman Amylase DS Assay 5. Boehringer Mannheim Pancreatic Amylase Method - Uses 2 monoclonal antibodies to inhibit >97% of the salivary isoenzymes activityMore reliable measure of pancreatic AMS than WGIM
• Boehringer Mannheim Pancreatic Amylase Method

  • Uses 2 monoclonal antibodies to inhibit >97% of the salivary isoenzymes activity. More reliable measure of pancreatic AMS than WGIM
• Boehringer Mannheim Pancreatic Amylase Method= More reliable measure of pancreatic AMS than WGIM
• Serum and Urine AMS- important in the diagnosis of acute pancreatitis
• serum AMS levels begin to rise 2-12 hours after onset of attack

  peak at 24 hours and return to normal levels within 3-5 days
• Acute pancreatitis:

  serum AMS levels begin to rise 2-12 hours after onset of attack peak at 24 hours and return to normal levels within 3-5 days Range from 250-1000 Somogyi units/dl (2.55 x ULN)
• 1. Perforated peptic ulcer, intestinal obstruction, cholecystitis, ruptured ectopic pregnanacy, mesenteric infarction, and acute appendicitis (inraabdominal diseases) usually <500 Somogyi units/dL
• • Other conditions: *makes elevated AMS a nonspecific finding

  1. Mumps and parotitis(salivary gland lesion)
  2. Perforated peptic ulcer, intestinal obstruction, cholecystitis, ruptured ectopic pregnanacy, mesenteric infarction, and acute appendicitis (inraabdominal diseases) usually <500 Somogyi units/dL
  3. Renal insufficiency/ failure
  4. Diabetic Ketoacidosis

  • Macroamylasemia
• Macroamylasemia

  • results when AMS molecule combines with immunoglobulins to form a complex that is too large to be filtered across the glomerulus
  • asymptomatic condition observed in approximately 1-2% of the population
  • reduction in normal renal clearance of the enzyme- thus, urinary excretion
  • its diagnostic significance lies in the need to differentiate it from other causes of hyperamylasemia 
• Specimen for AMS determination

  1.Serum

  2. Heparinized plasma

  3. Urine
• Heparinized plasma cannot use citrate,oxalate or EDTA
• Urine

  • timed-urine specimens for 1 to 2 hours- measures enzyme activity same manner as serum
  • taken at the same time as blood sample
  • if delayed: pH should be adjusted to neutral

  care must be taken to minimize bacterial growth

  > other specimen!

  1. peritoneal fluid and ascitic fluid (tumor)