(FINAL) CC1 LAB L4

Deamination

• Ammonia is produced during the __________ of amino acids in protein metabolism.

• It is removed from circulation and converted into urea in the liver (via ornithine cycle).

• Free ammonia is highly toxic to human cells.

• Despite its toxicity, ammonia is present in plasma at low concentrations.

Urea

• Ammonia is produced during the deamination of amino acids in protein metabolism.

• It is removed from circulation and converted into __________ in the liver (via ornithine cycle).

• Free ammonia is highly toxic to human cells.

• Despite its toxicity, ammonia is present in plasma at low concentrations.

Free ammonia

• Ammonia is produced during the deamination of amino acids in protein metabolism.

• It is removed from circulation and converted into urea in the liver (via ornithine cycle).

• __________ is highly toxic to human cells.

• Despite its toxicity, ammonia is present in plasma at low concentrations.

Low concentrations

• Ammonia is produced during the deamination of amino acids in protein metabolism.

• It is removed from circulation and converted into urea in the liver (via ornithine cycle).

• Free ammonia is highly toxic to human cells.

• Despite its toxicity, ammonia is present in plasma at __________.

Catabolism

Sources of ammonia:

• __________ of amino acids

• Bacterial metabolism in the intestinal lumen

• Anaerobic metabolism in skeletal muscles during exercise

Bacterial metabolism

Sources of ammonia:

• Catabolism of amino acids

• __________ in the intestinal lumen

• Anaerobic metabolism in skeletal muscles during exercise

Anaerobic metabolism

Sources of ammonia:

• Catabolism of amino acids

• Bacterial metabolism in the intestinal lumen

• __________ in skeletal muscles during exercise

Parenchymal cells

In the liver:

• Ammonia is taken up by __________

• Converted to urea via the Krebs-Henseleit (urea) cycle

• Urea is a non-toxic compound excreted in urine

Krebs-Henseleit (urea) cycle

In the liver:

• Ammonia is taken up by parenchymal cells

• Converted to urea via the __________

• Urea is a non-toxic compound excreted in urine

Urea

In the liver:

• Ammonia is taken up by parenchymal cells

• Converted to urea via the Krebs-Henseleit (urea) cycle

• __________ is a non-toxic compound excreted in urine

Ammonium ion (NH₄⁺)

In the blood:

• At normal physiologic pH, most ammonia exists as the __________

• There is a pH-dependent equilibrium between NH₃ and NH₄⁺

pH-dependent equilibrium

In the blood:

• At normal physiologic pH, most ammonia exists as the ammonium ion (NH₄⁺)

• There is a __________ between NH₃ and NH₄⁺

Ammonium ion (NH₄⁺)

In the kidneys:

• Ammonia is excreted as __________

• It helps buffer urine

Urine

In the kidneys:

• Ammonia is excreted as ammonium ion (NH₄⁺)

• It helps buffer __________

Hepatic failure, Reye’s syndrome

Blood ammonia measurement is useful in the following clinical conditions:

• __________

• ___________

• Inherited deficiencies of urea cycle enzymes

Hepatic failure

__________ (most common cause):

• Severe liver disease disrupts ammonia metabolism

• Blood ammonia levels may help determine prognosis

• The correlation between ammonia levels and hepatic encephalopathy is not always consistent

• Arterial ammonia is a better indicator of disease severity

Arterial ammonia

Hepatic failure (most common cause):

• Severe liver disease disrupts ammonia metabolism

• Blood ammonia levels may help determine prognosis

• The correlation between ammonia levels and hepatic encephalopathy is not always consistent

• __________ is a better indicator of disease severity

Reye’s syndrome

__________:

• Commonly occurs in children

• Serious and potentially fatal condition

• Often preceded by:

  • Viral infection

  • Aspirin use

• It is an acute metabolic disorder of the liver

• Autopsy findings: severe fatty infiltration of the liver

• Blood ammonia levels:

  • Correlate with disease severity and prognosis

  • Higher ammonia = worse prognosis

  • 100% survival if plasma NH₃ remains <5× normal levels

Unexplained nausea, vomiting, neurological deterioration

Inherited urea cycle enzyme deficiencies:

• Ammonia testing helps in diagnosis

• Suspect in neonates with:

  • __________

  • __________

  • __________, especially after feeding

• Other clinical uses:

  • Monitoring hyperalimentation (parenteral nutrition) therapy

  • Urine ammonia measurement:

    • Assesses the kidney's ability to produce ammonia

Hyperalimentation

Inherited urea cycle enzyme deficiencies:

• Ammonia testing helps in diagnosis

• Suspect in neonates with:

  • Unexplained nausea

  • Vomiting

  • Neurological deterioration, especially after feeding

• Other clinical uses:

  • Monitoring __________ (parenteral nutrition) therapy

  • Urine ammonia measurement:

    • Assesses the kidney's ability to produce ammonia

Accurate plasma ammonia results

Importance of specimen handling:

• Critical for __________

• Ammonia levels increase rapidly after collection due to in vitro amino acid deamination

In vitro amino acid deamination

Importance of specimen handling:

• Critical for accurate plasma ammonia results

• Ammonia levels increase rapidly after collection due to __________

Wet ice

Blood collection:

• Use venous blood

• Avoid trauma during collection

• Immediately place the specimen on __________

Heparin, EDTA

Anticoagulants:

• __________ is suitable

• __________ is suitable

Collection containers

__________:

• Must be checked for ammonia contamination/interference before use

0–4, 20

Processing of samples:

• Centrifuge at ___°C within ___ minutes of collection

• Separate plasma immediately after centrifugation

Freeze

Storage:

• Analyze as soon as possible

• If delayed, __________ the sample

• Frozen plasma is stable for several days at –20°C

–20°

Storage:

• Analyze as soon as possible

• If delayed, freeze the sample

• Frozen plasma is stable for several days at ___°C

2–3

Hemolysis:

• Must be avoided

• RBCs contain ___× more ammonia than plasma → can falsely increase results

Cigarette smoking

Patient preparation:

• Avoid __________ for several hours before collection

• __________ can increase ammonia levels (contamination source)

Increase

Substances that __________ ammonia levels:

• Ammonium salts, Asparaginase, Barbiturates, Diuretics, Ethanol, Hyperalimentation, Narcotic analgesics, other drugs

Decrease

Substances that __________ ammonia levels:

• Diphenhydramine, Lactobacillus acidophilus, Lactulose, Levodopa, and some Antibiotics

600

Analytical interference:

• Glucose >___ mg/dL (33 mmol/L) interferes with dry slide methods

Ion-selective electrode

Analytical Methods – Chemical Methods

• __________:

  • Diffusion of NH3 through a selective membrane into NH4Cl causes a pH change, which is measured potentiometrically

  • Good accuracy and precision; membrane stability may be a problem

Spectrophotometric

Analytical Methods – Chemical Methods

• __________:

  • NH3 + bromophenol blue → blue color

  • Measured using spectrophotometry

Glutamate dehydrogenase (GLDH)

Analytical Methods – Enzymatic Method

Catalyzed by __________:

• Enzymatic reaction of NH+, 2-oxoglutarate, and NADPH to form glutamate and NADP+, which is detected spectrophotometrically

• Most common on automated instruments; accurate and precise

NH+, 2-oxoglutarate, NADPH

Analytical Methods – Enzymatic Method

Catalyzed by Glutamate dehydrogenase (GLDH):

• Enzymatic reaction of __________, __________, and __________ to form glutamate and NADP+, which is detected spectrophotometrically

• Most common on automated instruments; accurate and precise

11–35

Reference Ranges:

• Adult Plasma:

  • Conventional Units: 19–60 μg/dL

  • SI Units: ______ μmol/L

• Adult Urine, 24 h:

  • Conventional Units: 140–1500 mg Nitrogen/d

  • SI Units: 10–107 mmol Nitrogen/d

• Child (10 d to 2 y) Plasma:

  • Conventional Units: 68–136 μg/dL

  • SI Unites: 40–80 μmol/L

10–107

Reference Ranges:

• Adult Plasma:

  • Conventional Units: 19–60 μg/dL

  • SI Units: 11–35 μmol/L

• Adult Urine, 24 h:

  • Conventional Units: 140–1500 mg Nitrogen/d

  • SI Units: ______ mmol Nitrogen/d

• Child (10 d to 2 y) Plasma:

  • Conventional Units: 68–136 μg/dL

  • SI Unites: 40–80 μmol/L

40–80

Reference Ranges:

• Adult Plasma:

  • Conventional Units: 19–60 μg/dL

  • SI Units: 11–35 μmol/L

• Adult Urine, 24 h:

  • Conventional Units: 140–1500 mg Nitrogen/d

  • SI Units: 10–107 mmol Nitrogen/d

• Child (10 d to 2 y) Plasma:

  • Conventional Units: 68–136 μg/dL

  • SI Unites: ______ μmol/L

Blood ammonia levels

In severe liver disease:

• Examples: cirrhosis, parenchymal liver damage

• Liver function is greatly impaired

• Ammonia is not properly removed from circulation

• __________ increase

Neurotoxic, astrocyte function

Effects of high ammonia levels:

• High NH₃ concentrations are __________

• Commonly associated with encephalopathy

• In the brain:

  • Alters metabolic processes

  • Leads to the accumulation of toxic substances

  • Impairs __________

Hyperammonemia

__________:

• Defined as elevated ammonia levels in the blood

• Associated with inherited deficiencies of urea cycle enzymes

Inherited metabolic disorders

Clinical significance:

• Measuring plasma ammonia is important for:

  • Diagnosis of __________

  • Monitoring these conditions

Frozen aliquots of human serum albumin

Recommended alternatives to serum-based controls:

• __________:

  • Containing known amounts of:

    • Ammonium chloride

    • Ammonium sulfate

• Commercial control solutions:

  • Contain known concentrations of ammonium sulfate

  • Readily available

Commercial control solutions

Recommended alternatives to serum-based controls:

• Frozen aliquots of human serum albumin:

  • Containing known amounts of:

    • Ammonium chloride

    • Ammonium sulfate

• __________:

  • Contain known concentrations of ammonium sulfate

  • Readily available

Serum-based controls

__________:

• Have unstable ammonia content

• Not reliable for accurate quality control