Alkaline Phosphatase ALP is affected by an individual's blood type (choose among ABO): 1. Increase in intestinal isoenzyme after a fatty meal 2. Decrease in intestinal isoenzyme after a fatty meal 3. Decrease in placental isoenzyme for pregnant women

1. B and O 2. A and AB 3. A and AB

(F) L1: Enzymes (Part 2) Flashcards by Kchianna Lauren Coronel

Major Clinical Enzymes: Phosphatases

Aka “Alkaline Orthophosphoric Monoester Phosphohydrolase”
Is non-specific
Is active in a pH of 10 to 15
It liberates inorganic phosphate with a byproduct of alcohol

Alkaline Phosphatase (ALP)

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Alkaline Phosphatase

What are the 5 major tissue sources of ALP?

Liver
Bone
Placenta
Intestine
Kidney

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Alkaline Phosphatase

Order the tissue sources from most abundant to least abundant:

Placenta
Intestine
Liver
Bone
Kidney

Liver (major source)
Bone (major source)
Placenta
Intestine
Kidney (minor source)

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Alkaline Phosphatase

An increase in bone isoenzyme is seen in what 2 age groups?

Growing children
Geriatric patients (>50 years old)

For children, since they are continuously undergoing bone resorption, bone tissues are continuously destroyed which liberates the enzymes

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Alkaline Phosphatase

An increase in ALP can also be seen in pregnant women how many weeks along in their pregnancy?

16-20 weeks

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Alkaline Phosphatase

ALP is affected by an individual’s blood type (choose among ABO):

Increase in intestinal isoenzyme after a fatty meal
Decrease in intestinal isoenzyme after a fatty meal
Decrease in placental isoenzyme for pregnant women

B and O
A and AB
A and AB

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Alkaline Phosphatase

What is the reference value?

30-90 U/L

Subjected to changes depending on multiple factors such as age, sex, race, and others

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Alkaline Phosphatase

What are the 4 major clinical isoenzymes?

Placental ALP
Intestinal ALP
Liver ALP
Bone ALP

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Alkaline Phosphatase (Specimen Considerations)

Match the elements with their functions:
1. One of the major components of ALP
2. An activator required in the analysis
3. An inhibitor

A. Phosphorus
B. Zinc
C. Magnesium

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Alkaline Phosphatase (Specimen Considerations)

In order to bind and inhibit phosphorus (since it is already a normal serum component), what buffer must be used?

2-amino-2-methyl-1-propanol (AMP)

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Alkaline Phosphatase (Specimen Considerations)

Does ALP increase or decrease after these activities/events?
- Hemolysis
- After ingestion of fatty meals (or food in general)
- Sample stored at 4ºC

Note: Fasting is not required for ALP testing

Increase

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Alkaline Phosphatase (Methods of Analysis)

This separates the different isoenzymes of ALP since each type exhibits different migrating patterns

Electrophoresis

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Alkaline Phosphatase (Electrophoresis)

What is/are the most anodal and least anodal isoenzymes?

Choose between: Liver, Bone, and Intestinal

Most anodal = Liver and Bone
Least anodal = Intestinal

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Alkaline Phosphatase (Electrophoresis)

What is Neuraminidase and Wheat Germ Lectin’s role in electrophoresis?

Separating agents for the separation of bone and liver ALP

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Alkaline Phosphatase (Electrophoresis)

High resolution electrophoresis using polyacrylamide gel with isoelectric focusing is used for what purpose?

To separate multiple overlapping ALP bands

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Alkaline Phosphatase (Methods of Analysis)

This exposes enzymes at 56ºC for 10-15 minutes wherein the most stable enzyme will persevere

Heat Fractionation/Stability Test

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Alkaline Phosphatase (Heat Fractionation/Stability Test)

Rank the ALPs from most heat stable to most heat labile?

Choices: Bone, Placenta, Intestinal, and Liver

Placental, Intestinal, Liver, Bone

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Alkaline Phosphatase (Methods of Analysis)

Uses chemical reagents at different concentrations to inhibit the different isoenzymes of ALP

Chemical Inhibition Test

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Alkaline Phosphatase (Chemical Inhibition Test)

Inhibits placental and intestinal ALP
Inhibits liver and bone ALP

A. Levamisole
B. Synthetic or 3M Urea
C. Phenylalanine

B and C
A

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Alkaline Phosphatase (Methods of Analysis)

The most specific method for ALP analysis (recommended by IFCC)
It measures however the total ALP amount and not the specific isoenzymes
It incorporates the concept of absolute specificity
A kinetic method that requires continuous monitoring at 405nm between a pH of 10 to 15
It reacts p-nitrophenylphosphate (PNPP) with ALP to produce p-nitrophenol and phosphate ions

Bowers and McComb (Szasz Modification)

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Diagnostic Significance of Alkaline Phosphatase

In cases of obstructive jaundice, ALP secretion in what organ is increased which leads to an increase in the total ALP?

Liver

When subjected under electrophoresis, the liver isoenzyme band may be increased

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Diagnostic Significance of Alkaline Phosphatase

In patients with Paget’s disease or osteotitis deformans, ALP secretion in what organ is increased?

Bone

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Diagnostic Significance of Alkaline Phosphatase

Patients who have undergone blood transfusion and cardiopulmonary bypass may exhibit a transient (increase/decrease) in ALP which will return to normal after some time

Decrease

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Diagnostic Significance of Alkaline Phosphatase

What ALP is increased in the ff. patients?
- Dialysis patients
- Patients with low bone mineral disease (BMD) of the hips
- Those with chronic kidney disease

B1x isoform (modified bone ALP)

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# Diagnostic Significance of Alkaline Phosphatase A prolonged (increase/decrease) in ALP is seen in patients with hypophosphatasia

Decrease

# Abnormal Forms of Alkaline Phosphatase - Seen in lung, breast, ovarian, and gynecological cancer - It is a co-migrator of bone ALP, wherein they are seen in the same place when subjected under electrophoresis - The most heat stable, surviving at 65ºC for 30 minutes - Is inhibited by phenylalanine

Regan ALP

# Abnormal Forms of Alkaline Phosphatase * Seen in patients with adenocarcinoma of the pancreas and bile duct, and pleural cancer * A variant of Regan ALP * It is inhibited by L-leucine and phenylalanine

Nagao ALP

# Abnormal Forms of Alkaline Phosphatase * Mirrors the placental ALP * Seen in serum and CSF specimens of patients with germ cell tumors * Determination of this is useful in differentiating pinealoma and germ cell tumor

Placental-like Alkaline Phosphatase (PLAP)

# Abnormal Forms of Alkaline Phosphatase Placental-like Alkaline Phosphatase (PLAP) is increased in patients with a (pinealoma/germ cell tumor)

Germ Cell Tumor

# Major Clinical Enzymes: Phosphatases - Aka “Acid Orthophosphoric Monoester Phosphohydrolase” - Catalyzes the same reaction with ALP except that this is acid-loving - Is maintained at a pH of 5 to 6 - Is mainly measured for male clinical chemistry, but not limited to it

Acid Phosphatase (ACP)

# Acid Phosphatase (ACP) An unknown specimen having ACP levels of greater than 50 U/L may be determined as having what fluid mixed in with the specimen?

Seminal fluid

# Acid Phosphatase (ACP) Arrange the tissue sources of ACP from most to least abundant: - Platelets - Liver - Prostate gland - RBCs - Bones

1. Prostate gland 2. RBCs 3. Platelets 4. Liver 5. Bones

# Acid Phosphatase (ACP) Reference Values for Males Determine if for Total ACP or Prostatic ACP: 1. 2.5 to 11.7 U/L 2. 0 to 3.5 ng/ml

1. Total ACP 2. Prostatic ACP

# Acid Phosphatase (ACP) Specimen Considerations 1. Sample should be free of hemolysis since it may lead to a falsely (increased/decreased) value of ACP as RBC can also be the source of the enzyme 2. A/an (increase/decrease) in the levels of ACP is seen when the sample is left at room temperature for 1 to 2 hours

1. Increased 2. Decrease

# Acid Phosphatase (ACP) Specimen Considerations This serves as the specific substrate for a quantitative endpoint reaction

Thymolphthalein monophosphate

# Acid Phosphatase (ACP) Specimen Considerations This substance may also be used when using an enzymatic kinetic method which requires continuous monitoring

A-naphthyl phosphate

# Acid Phosphatase (ACP) Specimen Considerations If the sample won’t be assayed immediately, it may be subjected to the following: 1. (Freezing/Refrigeration) 2. Acidification to a pH lower than 6.5 to make it stable for (two/three) days at room temperature

1. Freezing 2. Two (2)

# Acid Phosphatase (ACP) Specimen Considerations These serve as inhibitors for prostatic ACP

20mmol of L-tartrate ions

# Acid Phosphatase (ACP) Specimen Considerations This serves as an inhibitor for RBC ACP

1mmol of cupric sulfate + 2% formaldehyde

# Acid Phosphatase (ACP) Diagnostic Significance ACP is mainly used in males for the detection of what? Note: The number of ACP increases due to an increase in the number of cells when diagnosed with this

Prostatic adenocarcinoma

# Acid Phosphatase (ACP) Diagnostic Significance ACP levels fall (faster/slower) than the prostate specific antigen (PSA) in cases of surgical treatment

Faster | PSA is mainly concerned with immunology rather than CC

# Acid Phosphatase (ACP) Diagnostic Significance ACP determination is also useful for rape cases wherein vaginal washings are examined for seminal fluid ACP which can persist for up to how many days?

4 days

# Acid Phosphatase (ACP) Diagnostic Significance This is a form of ACP that is resistant to L-tartrate and a known inhibitor of prostatic ACP, which is seen in chronic and hairy cell leukemia

Tartrate Resistant Acid Phosphatase (TRAP)

# Acid Phosphatase (ACP) Diagnostic Significance Increased or Decreased ACP? - Urinary tract obstruction - Acute urinary retention - Extensive prostatic massage - Prostatic inflammation - Infarction or Ischemia - Prostatic manipulations (e.g. needle biopsy and cytoscopy) - Prostatic carcinoma - Breast, lung, and thyroid carcinoma - Gaucher’s disease - Niemann Pick disease - Thrombocytopenia

Increased

# Major Clinical Enzymes: Transferases/Transaminases Aspartate and α-ketoacid reacts with this enzyme which will catalyze the transfer of an amino group to produce oxaloacetate and glutamate

Aspartate Aminotransferases or Serum Glutamic Oxaloacetic Transaminase (AST/SGOT)

# Major Clinical Enzymes: AST/SGOT What are the 3 major tissue sources of AST?

1. Cardiac (major) 2. Liver 3. Skeletal (minor) | Other sources: Kidney, Pancreas, and RBCs

# Major Clinical Enzymes: AST/SGOT What is the reference value?

5 to 37 U/L

# Major Clinical Enzymes: AST/SGOT What are the 2 isoenzymes?

1. Cytoplasmic AST (more predominant) 2. Mitochondrial AST

# AST/SGOT Methods of Analysis - This method maintains a pH of 7.5 - It pairs AST with malate dehydrogenase (MD) to monitor changes in absorbance at 340nm - AST reacts with aspartate and a-ketoglutarate to produce oxaloacetate with NADH - Oxaloacetate will be acted upon by MD to produce malate and NAD

Karmen Method

# AST/SGOT Diagnostic Significance AST is beneficial in the evaluation of (acute/chronic) myocardial infarction as well as hepatocellular disorders and skeletal muscle involvement

Acute

# AST/SGOT Diagnostic Significance In cases of Acute MI, AST is a major enzyme used to determine the severity of the heart attack wherein AST levels should exhibit the following: 1. A (rise/fall) in values within 6 to 8 hours after infarction 2. A (peak/drop) at 24 hours after infarction 3. A/an (increase/decrease) back to normal within 5 days after infarction

1. Rise 2. Peak 3. Decrease

# AST/SGOT Diagnostic Significance Match the ff. timeframes with the events during an Acute MI: 1. A rise in value within (blank) after infarction 2. A peak at (blank) after infarction 3. A decrease back to normal within (blank) after infarction A. 24 hours B. 5 days C. 6 to 8 hours

1. C 2. A 3. B

# AST/SGOT Diagnostic Significance As a physician, there are two things to be considered when diagnosing AMI: 1. Specificity of the (enzyme/substrate) 2. The (rise/fall)

1. Enzyme 2. Rise

# AST/SGOT Diagnostic Significance TOF: AST is specific for diagnosing AMI | AMI: Acute Myocardial Infarction

False (it can be sourced from other organs)

# AST/SGOT Diagnostic Significance TOF: Monitoring and waiting for the rise of AST during an emergency AMI case is beneficial

False (it takes 6 to 8 hours before levels start to increase, and time is key in diagnosing AMI, a life-threatening disease)

# AST/SGOT Diagnostic Significance AST is also used in monitoring therapy for what kind of drugs which are able to increase AST levels 3x than the upper limits of the reference value?

Hepatotoxic drugs

# AST/SGOT Diagnostic Significance Chronic disorders result to a significant (increase/decrease) in AST level

Increase

# Major Clinical Enzymes: Transferases/Transaminases - Alanine and α-ketoglutarate reacts with this enzyme to produce glutamate and pyruvate - Is significant in the evaluation of hepatic disorders - It can also be used to monitor the course of liver treatment and effects of drug therapy

Alanine Aminotransferase or Serum Glutamic Pyruvic Transaminase (ALT/SGPT)

# Major Clinical Enzymes: Transferases/Transaminases Which among AST and ALT is a more specific and sensitive screening test for post transfusion hepatitis and occupational toxic exposure?

Alanine Aminotransferase or Serum Glutamic Pyruvic Transaminase (ALT/SGPT)

# ALT/SGPT Specimen Considerations What 4 samples can be used for ALT analysis?

1. Plasma 2. Bile 3. CSF 4. Saliva

# ALT/SGPT Specimen Considerations ALT analysis requires what coenzyme?

Pyridoxal phosphate (vitamin B6)

# ALT/SGPT Specimen Considerations A hemolyzed specimen may falsely increase the levels of ALT up to how many times?

10x

# ALT/SGPT Specimen Considerations What anticoagulant tube must not be used for ALT analysis considering that it inhibits the activity of the enzyme?

Heparin (green-top)

# ALT/SGPT Methods of Analysis - This method maintains a pH of 7.5 and is read at 340nm - This couples ALT with lactate dehydrogenase (LD) which produces pyruvate - Pyruvate when combined with NADH and an H ion will be acted upon by LD to produce lactate and NAD

Coupled Enzyme Reaction

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: Major organ is the HEART

AST/SGOT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: Major organ is the LIVER

ALT/SGPT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: Substrate is Aspartic a-ketoglutaric acid

AST/SGOT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: Substrate is Alanine a-ketoglutaric acid

ALT/SGPT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: End products are glutamic acid and oxaloacetic acid

AST/SGOT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: End products are glutamic acid and pyruvic acid

ALT/SGPT

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: The color developer is 2,4 DNPH

BOTH | Note: The developer is for spectrophotometer

# Transferases & Transaminases (Comparison Table) Identify if AST/SGOT or ALT/SGPT: The color intensifier is 0.4N of NaOH

BOTH

# Transferases & Transaminases (Comparison) Identify if AST/SGOT or ALT/SGPT: Disease involves more of the muscle

AST/SGOT

# Transferases & Transaminases (Comparison) Identify if AST/SGOT or ALT/SGPT: Disease is more localized in the liver

ALT/SGPT

# Transferases & Transaminases (Comparison) In viral hepatitis, both ALT and AST levels are elevated. In order to determine what enzyme to use, what ratio is needed?

De Ritis Ratio (ratio between ALT and AST)

# Transferases & Transaminases (Comparison) If the De Ritis Ratio is greater than 1.0 which is commonly seen in acute hepatitis, what enzyme may be used?

ALT/SGPT

# Major Clinical Enzymes - Aka “Alpha-1-4 glucan-4-glucohydrolase” - It is a standalone enzyme that catalyzes a specific reaction - Starch and glycogen will be acted upon by this enzyme to produce a reducing sugar (glucose) - It is the smallest enzyme (MW: 50,000-55,000 Daltons) hence is freely filtered by the glomerulus - Also one of the earliest pancreatic markers

Amylase (AMS)

# Amylase (AMS) What are the 2 major tissue sources?

1. Acinar cells of the pancreas 2. Salivary glands | Other sources: Adipose, Fallopian tubes, SI, and Skeletal muscles

# Amylase (AMS) What are the 2 reference values?

1. 60 to 180 Somogyi units per dL (SU/dL) 2. 95 to 290 U/L

# Amylase (AMS) What are the 2 isoenzymes?

1. Salivary (S) type/ Ptyalin 2. Pancreatic (P) type/ Amylopsin

# Amylase (AMS) This isoenzyme is inhibited by wheat germ lectin

Salivary (S) type/ Ptyalin

# Amylase (AMS) Specimen Considerations Give 3 interferences of AMS

1. Heparin 2. TAGs 3. Wheat germ

# Amylase (AMS) Specimen Considerations An increase in AMS activity will be solved by (diluting/concentrating) the specimen

Diluting

# Amylase (AMS) Specimen Considerations A/an (increase/decrease) in AMS levels can be caused by the use of morphine and other opiates

Increase

# Amylase (AMS) Specimen Considerations What is the general substrate used for AMS?

Starch

# Amylase (AMS) Methods of Analysis - The reference method reported in Somogyi units (SU) - Starch is broken down by AMS to produce glucose which will then be subjected to other glucose methods in order to be measured - Amount of glucose = amount of amylase

Saccharogenic

# Amylase (AMS) Methods of Analysis This measures enzyme activity by the decrease in substrate concentration

Amyloclastic

# Amylase (AMS) Methods of Analysis This measures enzyme activity by the increase of color intensity of a soluble dye-substrate solution

Chromogenic

# Amylase (AMS) Methods of Analysis - This is a kinetic method which requires a continuous monitoring technique - Amylase is coupled with a number of enzymes such as glucosidase, hexokinase, and G6PD - Maltopentose is acted upon by AMS to produce maltotriose and maltose - Maltotriose and maltose will be acted upon by glucosidase to produce 5-glucose which will be combined with ATP to be acted upon by hexokinase - 5-glucose-6-phosphate along with ADP will be combined with NAD to be acted upon by G6PD to produce 5,6-phosphogluconolactone

Coupled Enzyme

# Amylase (AMS) Methods of Analysis This is used to determine the state of renal filtration, and as amylase is freely filtered in the glomeruli, using their ratio may determine the state of renal filtration

Creatinine (for the amylase/creatinine ratio)

# Amylase/Creatinine Ratio What is the reference value?

1% to 4% (or 0.01 to 0.04)

# Amylase/Creatinine Ratio For patients with acute pancreatitis, the ratio may reach what values?

4% to 15%

An increase in the AMS levels might indicate a/an (increase/decrease) in the filtration on this enzyme, which is an indication of an impaired glomerular function

Increase

# Amylase (AMS) Diagnostic Significance AMS determination is useful in the evaluation of acute pancreatitis and parotitis since it can also be sourced from where?

Salivary glands

# Amylase (AMS) Diagnostic Significance In cases of both pancreatitis and parotitis, an increase in plasma AMS may be seen along with an increase of AMS in the urine for up to how many days?

7 days

# Amylase (AMS) Diagnostic Significance AMS may be used to evaluate the presence and/or severity of acute pancreatitis wherein the levels of the enzyme may exhibit the following: 1. A (rise/fall) in 2 to 12 hours 2. A (peak/drop) in 24 hours 3. A/an (decrease/increase) back to normal levels in 3 to 5 days

1. Rise 2. Peak 3. Decrease

# Amylase (AMS) Diagnostic Significance Match the ff. timeframes with the events during acute pancreatitis: 1. A rise in value within (blank) 2. A peak at (blank) 3. A decrease back to normal within (blank) A. 2 to 12 hours B. 24 hours C. 3 to 5 days

1. A 2. B 3. C

# Amylase (AMS) Diagnostic Significance TOF: In cases of renal failure, an increase in AMS plasma will be seen along with a decrease of AMS in the urine

True

# Amylase (AMS) Diagnostic Significance This is a form of amylase combined with an immunoglobulin, wherein it cannot be filtered by the glomerulus due to its larger size

Macroamylasemia

# Amylase (AMS) Diagnostic Significance Increased or Decreased Amylase? - Acute pancreatitis - Ectopic pregnancy - Peptic ulcer - Alcoholism - Mumps

Increased

# Major Clinical Enzymes - Aka “Triacylglycerol acylhydrolase” - It catalyzes the degradation of fats and triacylglycerols (TAG) which will result to the formation of alcohol with fatty acids, and 2-monoglyceride intermediates with fatty acids, respectively - It hydrolyzes the ester linkages of fats

Lipase (LPS)

# Lipase (LPS) Lipase is the most specific pancreatic marker however one disadvantage is that?

It is a LATE pancreatic marker

# Lipase (LPS) What is the major tissue source of lipase?

Pancreas

# Lipase (LPS) What is the reference value?

Between 0 to 1 U/mL

# Lipase (LPS) Specimen Considerations What is the main substrate?

Olive oil (Triolein) | Note: It is the purest form of TAG

# Lipase (LPS) Specimen Considerations What is the coenzyme needed for the reaction to work?

Colipase (secreted by the pancreas together with bile salts)

# Lipase (LPS) Specimen Considerations Lipase is inhibited by what component from red blood cells?

Hemoglobin

# Lipase (LPS) Methods of Analysis The most commonly used method of analysis however it does not use 50% olive oil

Peroxidase Coupling

# Lipase (LPS) Methods of Analysis - The reference method for LPS analysis - Involves the hydrolysis of olive oil after 24 hours at 37 ºC and the titration of fatty acids using NaOH - The end product is fatty acids - The application of titration makes the method less efficient since it can be time-consuming and labor-intensive

Cherry Crandal Method

# Lipase (LPS) Methods of Analysis What are the 3 methods of analysis?

1. Peroxidase Coupling 2. Tietz and Fiereck 3. Cherry Crandal Method

# Lipase (LPS) Diagnostic Significance In cases of chronic pancreatitis, acinar cell degradation is exhibited resulting to a/an (increase/decrease) in amylase along with lipase production

Decrease

# Lipase (LPS) Diagnostic Significance Determination of LPS may also be used in diagnosing the presence and/or severity of acute pancreatitis. Wherein the LPS levels will exhibit the following: 1. A (rise/fall) in 6 hours 2. A (peak/drop) in 24 hours 3. A/an (increase/decrease) to normal levels in 8 to 14 days

1. Rise 2. Peak 3. Decrease

# Lipase (LPS) Diagnostic Significance Match the ff. timeframes with the events during acute pancreatitis: 1. A rise in value within (blank) 2. A peak at (blank) 3. A decrease back to normal within (blank) A. 24 hours B. 6 hours C. 8 to 14 days

1. B 2. A 3. C

What are the 5 function markers of the pancreas?

1. Amylase 2. Lipase 3. Trypsin 4. Chymotrypsin 5. Elastase-1

Familiarize with the Amylase/Creatinine Ratio Formula

(urine amylase/serum amylase) x (serum creatinine/urine creatinine) x 100

(F) L1: Enzymes (Part 2) Flashcards

(115 cards)