Order the NPNs from most significant to least (in %) - Amino acids - Ammonia - Creatine - Creatinine - Urea - Uric acids I will give the percentages in the answer, just familiarize there

1. Urea (45%) 2. Amino Acids (20%) 3. Uric Acids (20%) 4. Creatinine (5%) 5. Creatine (1-2%) 6. Ammonia (0-2%)

(M) Lec 3: Non-Nitrogenous Proteins Flashcards by Kchianna Lauren Coronel

these have low molecular weight
they contain nitrogen
are distinguished from proteins based on their size
are metabolic by-products/waste of proteins, nucleic acids, and muscle metabolism

Non-Protein Nitrogenous Compounds (NPNs)

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A high NPN value is (advantageous or disadvantageous) to man

Disadvantageous

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In what excretions (2) are a large number of NPNs found in?

Urine and Stool

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In what body fluid are NPNs ideally found in small amounts/minimal concentrations?

Blood

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What organ is responsible for properly excreting NPNs?

Kidney

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NPNs are converted into proteins by bacteria found in the stomachs of (animals or humans)

Animals

Animals further convert NPNs into proteins while NPNs are the end products in humans (we lack the bacteria that convert it)

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Where are NPNs incorporated in to increase its crude protein value which is measured based on nitrogen content?

Animal feeds

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Crude protein values of NPNs in animal feeds

Protein: % N?
Urea: % N?
Melamine: % N?

Protein: 16%
Urea: 47%
Melamine: 66%

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Order the NPNs from most significant to least (in %)

Amino acids
Ammonia
Creatine
Creatinine
Urea
Uric acids

I will give the percentages in the answer, just familiarize there

Urea (45%)
Amino Acids (20%)
Uric Acids (20%)
Creatinine (5%)
Creatine (1-2%)
Ammonia (0-2%)

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What 4 NPNs are part of the parameters for a kidney function test?

Urea
Amino acids
Creatinine
Creatine

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What NPN is part of the parameters for a liver function test?

Ammonia

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What NPN will significantly increase if there is an increased result for NPNs in general?

Urea (highest concentration)

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This NPN is technically not yet part of the “NPNs classified as waste products” as it is “not really” a protein yet

Amino acids

Note: From protein > amino acids > urea

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Which among creatinine and creatine is the end product?

Creatinine (creatine signifies incomplete muscle metabolism)

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Clinically Significant NPNs

This is the major excretory product of oxidative protein and amino acid metabolism

Urea or BUN

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Clinically Significant NPNs

This is synthesized in the liver from CO2 and ammonia that arises from the deamination of amino acids

Urea or BUN

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The synthesis of UREA in the liver from CO2 and ammonia that arises from the deamination of amino acids is known as what cycle?

Clue: It has 3 names

Urea cycle
Ornithine cycle
Kreb’s Henseleit cycle

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Urea is excreted in both urine and stool, but what percentage of it actually makes it to the urine?

90%

Consists of half of the urinary solids

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Determination of Blood Concentration (UREA)

Abnormal levels of urea can give an idea about an individual’s renal function and perfusion, does it indicate acute or chronic disorders?

BOTH

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Determination of Blood Concentration (UREA)

A diet rich in this component will result to high urea formation

High protein diet

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Determination of Blood Concentration (UREA)

Urea is a good indicator for an individual’s state of what?

Hydration

Normal Urea = Hydrated
Abnormal Urea = Possible dehydration

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Determination of Blood Concentration (UREA)

Refers to how FAST the body is able to catabolize proteins

Protein Catabolism Rate

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Determination of Blood Concentration (UREA)

Urea is a good indicator for an individual’s intake of what?

Nitrogen

Urea has the HIGHEST nitrogen concentration among the NPNs

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Urea Determination

This has been used to refer to a MEASUREMENT OF UREA, not the actual urea

Blood Urea Nitrogen (BUN)

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# Urea Determination Actual Urea has a formula of what? | Actual urea is the level of urea minus the nitrogen

BUN x 2.14 mg/dL | The CF for urea is 2.14

# Urea Determination What is the conversion factor for actual urea when you have computed for the conventional unit (mg/dL) and would like to report it as an SI unit (mmol/L)? | Simply multiply the obtained mg/dL value with this CF to get mmol/L

0.357 or 0.36 mmol/L

# Analytical Methods in Determining Urea Which among enzymatic and chemical reactions are the direct and indirect methods for determining urea?

Enzymatic - indirect Chemical - direct

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - most common enzymatic method - couples the urease with glutamate dehydrogenase (2 reactions) - the amount of NAD produced corresponds to the amount of urea

Coupled Urease (Glutamate Dehydrogenase) Method

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) In Coupled Urease (Glutamate Dehydrogenase) Method, what are the end products [2] after the coupled reactions?

Glutamate and H2O

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) In Coupled Urease (Glutamate Dehydrogenase) Method, the amount of glutamate and H2O is not the main analyte being measured, instead what it is?

Nicotinamide Adenine Dinucleotide (NAD)

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - ammonia will result in a color change with the help of a pH indicator - thiosemicarbazide and ferric ions are added to enhance color development - color intensity corresponds to the amount of urea | Note: W/o the reagents in the 2nd bullet, the color will be very light

Indicator Dye

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - urea is broken down into ammonia and bicarbonate which produces conductive activity - amount of conductivity produced is correspondent to the amount of urea | Note: This method has low sensitivity

Conductimetric

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - this measures the production of indophenol blue when phenol is added as a reagent - ammonia will result in a color change with the help of a pH indicator

Berthelot Reaction

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - urea will produce ammonia and CO2 (the ammonia can be measured by treating it with Berthelot reagents) - is commonly and routinely done

Hydrolysis of Urea (by urease)

# Analytical Methods in Determining Urea (Enzymatic/Indirect Method) - this is the REFERENCE METHOD for measuring urea - the GOLD STANDARD

Isotope Dilution Mass Spectrometry (IDMS) ## Footnote Note: This is not only for urea, it is also the reference method for other tests

# Analytical Methods in Determining Urea (Chemical/Direct Method) - most commonly used direct method - diacetyl is added to urea to produce diazine - it appears as a yellow-colored compound - it is measured using a spectrophotometer and it has a higher sensitivity than other methods

Diacetyl Monoxime (DAM)

# Analytical Methods in Determining Urea (Chemical/Direct Method) In the Diacetyl Monoxime (DAM) Method, the production of diazine from the combination of urea and diacetyl is known as what reaction?

Fearon's Reaction

# Analytical Methods in Determining Urea (Chemical/Direct Method) - the addition of o-phthaldehyde to urea will produce isoindoline which is acted upon by N-(1-naphthyl)-ethylenediamine resulting in a colored product - the resulting color is dependent on the indicator added

O-phthaldehyde Method

# Specimen Requirements (Urea) What 3 bodily fluids can be used?

1. Serum 2. Plasma 3. Urine

# Specimen Requirements (Urea) Urea should not be contamined with: 1. Fluoride 2. Citrate 3. Ammonia Which among these results in false increased and false decreased values? | Note: I will explain the rationale in the results

Falsely decreased: Fluoride and Citrate Falsely increased: Ammonia ## Footnote Fluoride and Citrate - inhibits urease Ammonia - a fellow NPN

# Specimen Requirements (Urea) TOF: You must fast before submitting a sample for urea testing

False (a non-fasting sample is needed)

# Reference Values for Urea - For serum? - For 24H urine?

Serum: 6-20 mg/dL Urine: 12-20 g/day

# Disease Correlations of UREA - elevated levels of all nitrogenous substances (urea has the most striking prominence) - it has three types depending on the organ of origin

Azotemia

# Disease Correlations of UREA What are the 3 kinds of azotemia?

1. Pre-renal 2. Renal 3. Post-renal

# Disease Correlations of UREA - accumulation of NPNs in the blood due to the inability of urea to enter the kidneys (before it even arrives at the kidneys) - caused by conditions already existing in the blood (specifically a decrease in renal perfusion) - caused by dehydration, shock, and congestive heart failure

Pre-renal Azotemia

# Disease Correlations of UREA In Pre-renal Azotemia, there is a decreased rate of what but with normal renal function?

Glomerular Filtration Rate (GFR)

# Disease Correlations of UREA In Pre-renal Azotemia, increased urea comes with (decreased, normal, or increased) blood creatinine

Normal

# Disease Correlations of UREA - the TRUE kidney disease - there is increased BUN due to poor excretion - caused by either acute or chronic kidney disease and glomerulonephritis - can cause coma and neuropsychiatric changes

Renal Azotemia

# Disease Correlations of UREA In Renal Azotemia, striking urea levels come with (decreased, normal, or increased) blood creatinine

Increased (slowly rising)

# Disease Correlations of UREA Renal Azotemia Criteria (give the values for the ff. analytes:) 1. BUN 2. Creatinine 3. Uric Acid

1. BUN - ≥100 mg/dL 2. Creatinine - ≥20 mg/dL 3. Uric Acid - ≥12 mg/dL

# Disease Correlations of UREA - there is marked elevation in plasma urea accompanied by acidemia and electrolyte imbalance (potassium is elevated) - the kidneys fail to eliminate waste products - characterized by anemia (normocytic normochromic), uremic frost (dirty skin of metallic color), edema, foul breath, and urine in sweat

Uremia | Note: Only BUN is high in this disease (because it is found in plasma)

# Disease Correlations of UREA What are the normal values of the BUN to CREATININE (B:C) ratio which differentiates the causes of abnormal urea in the blood?

10:1 to 20:1

# Disease Correlations of UREA Indicate if: A. Low B:C Ratio (< 10:1) B. Increased B:C w/ normal creatinine C. Increased B:C w/ high creatinine - decreased urea production - low-protein diet - acute tubular necrosis - repeated dialysis - hepatic disease

# Disease Correlations of UREA Indicate if: A. Low B:C Ratio (< 10:1) B. Increased B:C w/ normal creatinine C. Increased B:C w/ high creatinine - pre-renal azotemia - high-protein intake - thyrotoxicosis - GI hemorhhage - dehydration - catabolic states

# Disease Correlations of UREA Indicate if: A. Low B:C Ratio (< 10:1) B. Increased B:C w/ normal creatinine C. Increased B:C with high creatinine - post-renal azotemia - pre-renal azotemia with renal disease - uremia - renal failure

TOF: Urea is the hardest waste to remove in dialysis due to its abundance

False (the easiest)

# Answer if increased or decreased BUN - azotemia (all three types) - fever - stress - burns - high-protein diet - dehydration - chronic renal disease

Increased

# Answer if increased or decreased BUN - poor nutrition - high fluid intake (overhydration) - pregnancy - severe liver disease - hormonal effects - malnutrition - hepatic diseases

Decreased

- end product of muscle metabolism - is not reabsorbed and minimally secreted in the kidneys - amount generated is proportional to the muscle mass - is not affected by protein intake - a marker for glomerular filtration rate which makes it an index for overall renal function - measures the completeness of 24H urine

Creatinine

Once creatine is produced via muscle metabolism, it will be (dehydrated, hydrolyzed, methylated, or deaminated) to form creatinine

Dehydrated

Creatinine is also produced by 3 amino acids, what are those?

Methionine, Arginine, and Lysine

TOF: Creatinine is not easily removed via dialysis

True

# Increased or Decreased Creatinine? - impaired renal function - chronic nephritis - muscular diseases - congestive heart failure

Increased

# Increased or Decreased Creatinine? - decreased muscle mass - advanced and severe lung disease - pregnancy - inadequate dietary protein

Decreased

# Analytical Methods in Determining Creatinine - most frequently used and first described in 1886 - creatinine reacts with picric acid in an alkaline solution to form a red-orange tautomer of creatinine picrate - has many interferences (ascorbate, glucose, uric acid, and a-ketoacids)

Direct Jaffe Reaction

# Analytical Methods in Determining Creatinine This is a saturated reagent comprised of picric acid and 10% NaOH

Jaffe Reagent

# Analytical Methods in Determining Creatinine - requires automated equipment to measure the rate of change of absorbance - it takes more than one measurement - rate of color production is measured (spectrophotometer)

Kinetic Jaffe Method or Janovsky-like Reaction

# Analytical Methods in Determining Creatinine - uses creatininase or creatinine amidohydrolase, creatine kinase, pyruvate kinase, and lactate dehydrogenase - peroxidase may also be used in a dry slide technique - this eliminates the non-specificity of the Jaffe reaction as it is more specific

Enzymatic Method

# Analytical Methods in Determining Creatinine (Enzymatic Methods) - produces lactate with NAD (which are measured) as these are always directly proportional to creatinine

Creatininase-Creatine Kinase

# Analytical Methods in Determining Creatinine (Enzymatic Methods) - using peroxidase, the end product exhibits a color through the addition of an indicator to measure the color intensity

Creatininase-Hydrogen Peroxide

# Analytical Methods in Determining Creatinine - modified Jaffe reaction with adsorbent (done via elution) - the picric acid with adsorbent now functions to remove interferences

Chemical Method

# Analytical Methods in Determining Creatinine What adsorbent is composed of sodium aluminum silicate?

Lloyd's reagent

# Analytical Methods in Determining Creatinine What adsorbent is composed of aluminum magnesium silicate?

Fuller's earth

# Analytical Methods in Determining Creatinine The reference method for creatinine measurement

Isotope Dilution Mass Spectrometry (IDMS)

# Specimen Requirements for Creatinine What 3 body fluids can be used?

1. Plasma 2. Serum 3. Urine

# Specimen Requirements for Creatinine Creatinine requires a (fasting/non-fasting) sample

Non-fasting

TOF: Protein ingestion cannot elevate creatinine levels

False (it can transiently elevate but only minimally)

# Interferences of Creatinine (falsely increased or decreased) - a-ketoacids - glucose - ascorbic acids - uric acid - cephalosporin - high ketones

False increase

# Interferences of Creatinine (falsely increased or decreased) - bilirubin - hemoglobin (hemolyzed sample) - contamination

False decrease

# Creatinine Interferences Bilirubin and hemoglobin cause what resulting in a false decrease in results?

Dilution

# Creatinine Interferences This is an anesthesia that can cause interferences in enzymatic methods

Lidocaine

# Creatinine Interferences This can cause interferences in BOTH enzymatic and Jaffe reactions

Dopamine

# Creatinine Interferences This causes false increased levels using Jaffe's reaction

Cephalosporin

# Reference Values for Creatinine Choices: A. Female adult (Jaffe reaction) B. Female adult (Enzymatic method) C. Male adult (Jaffe reaction) D. Male adult (Enzymatic method) E. Children (Jaffe reaction) F. Children (Enzymatic method) Given: 0.6 - 1.1

A and D

Creatinine has different reference values because it depends on the what of the individual?

Muscle Mass

What is the conversion factor of creatinine to its SI unit?

88.7 (to umol/L)

# Creatinine 24H Urine Values Male or Female: 800-2000 mg/dL

Male

# Creatinine 24H Urine Values Male or Female: 600-800 mg/dL

Female

# Disease Correlations of Creatinine Elevated creatinine in the blood is correlated to what kidney disease?

Renal Failure

# Disease Correlations of Creatinine Creatinine is used in what test to measure glomerular filtration rate and how effective the kidneys are in filtering wastes in the blood

Clearance Test

- major end product of purines (adenine and guanine) metabolism - the final breakdown of nucleic acid catabolism in humans - is freely filtered but significantly reabsorbed and secreted in the tubules

Uric Acid

For higher primates, uric acid is the end product of purine metabolism but in some animals, uric acid is further converted into what?

Allantoin

# Uric Acid More than (>) 95% of uric acid exists as what kind of urate at a pH of 7.4 (slightly alkaline)?

Monosodium Urate

# Uric Acid Uric acid at a pH of 5.4 and below exists as?

Uric acid (lol) | Note: It has a different form when in an alkaline environment

# Uric Acid A concentration of > 6.4 mg/dL in plasma is considered (saturated, normal, unsaturated)

Saturated

# Hyperuricemia or Hypouricemia - leukemia - lymphoma - multiple myeloma - drugs, diet, and obesity - hypertriglyceridemia - renal disease - gout - Lesch Nyhan syndrome - ethanol consumption

Hyperuricemia

# Hyperuricemia or Hypouricemia - Fanconi's - Wilson's - Hodgkin's - Bronchogenic CA - Xanthinuria

Hypouricemia

# Analytical Methods in Determining Uric Acid - the simplest and most specific method - is acted upon by uricase to produce allantoin

Enzymatic Method

# Analytical Methods in Determining Uric Acid (Enzymatic Methods) - measures the difference in absorbance before and after adding uricase - a decrease in absorbance is proportional to the uric acid content

Spectrophotometric

# Analytical Methods in Determining Uric Acid (Enzymatic Methods) Uric acid is measured at a peak absorbance of what using the spectrophotometric method?

293nm

# Analytical Methods in Determining Uric Acid (Enzymatic Methods) Which among the two is the Coupled Enzymatic Reaction I and II? 1. is coupled with catalase to produce a colored compound 2. is coupled with peroxidase to produce a colored compound

1. CATALASE = enzymatic reaction I 2. PEROXIDASE = enzymatic reaction II

# Analytical Methods in Determining Uric Acid (Chemical Methods) - involves a reaction with uric acid and alkaline phosphogluconate with tungsten blue as the end product - the principle is a redox reaction upon incubation

Phosphotungstic Method

# Analytical Methods in Determining Uric Acid (Chemical Methods) In the spectrophotometric method, this additive is used in Brown's, Benedict's, Folin's, and Newton's methods

Sodium Bicarbonate

# Analytical Methods in Determining Uric Acid (Chemical Methods) In the spectrophotometric method, this additive is used in Archibal's, Caraway's, and Henry's methods

Sodium Cyanide

# Analytical Methods in Determining Uric Acid (Chemical Methods) In spectrophotometry, this will occur after the addition of the additives to inactivate non-uric acid reactants to isolate uric acid for measurement

Lag Phase

# Analytical Methods in Determining Uric Acid (Chemical Methods) The reference method for uric acid measurement

Isotope Dilution Mass Spectrometry (IDMS)

# Specimen Requirements for Uric Acid TOF: Uric acid is stable in both serum and plasma for 3 days at room temperature

False (serum and URINE, not plasma) | The other parameters are correct

# Specimen Requirements for Uric Acid This substance cannot be used as an anticoagulant as it can interfere with results

Potassium oxalate

# Specimen Requirements for Uric Acid What are the 2 major interferences that falsely decrease results?

1. Ascorbic Acid 2. Bilirubin

# Reference Values for Uric Acid For females?

2.6 - 6.0 mg/dL

# Reference Values for Uric Acid For males?

0.5 - 7.2 mg/dL

# Reference Values for Uric Acid What is the conversion factor to convert mg/dL to mmol/L?

0.0595

# Disease Correlations for Uric Acid (hyperuricemia) - pain and inflammation in the joints - crystals are found in the synovial fluid - can be treated with allopurinol

Gout

# Disease Correlations for Uric Acid (hyperuricemia) - leukemia, lymphoma, multiple myeloma, and polycythemia vera - can be treated with allopurinol

Increased nuclear metabolism

# Disease Correlations for Uric Acid (hyperuricemia) - due to increased GFR and tubular secretion

Chronic renal disease

# Disease Correlations for Uric Acid (hyperuricemia) - an inborn error of purine metabolism (X-linked) - decreased hypoxanthine-guanine phosphoribosyltransferase (HGPRT)

Lesch-Nyhan Syndrome (aka Juvenile Gout)

# Disease Correlations for Uric Acid (hyperuricemia) - glycogen storage diseases - toxemia of pregnancy and lactic acidosis - due to increased dietary intake and ethanol consumption

Other causes of hyperuricemia (secondary to the disease)

# Disease Correlations for Uric Acid - Fanconi's syndrome (congenital tubular necrosis) - Wilson's disease - Hodgkin's disease

Hypouricemia

What NPNs can be measured with the IDMS method?

1. Urea 2. Creatinine 3. Uric Acid

- a byproduct of protein catabolism that arises from the deamination of amino acids due to the action of intestinal flora - also released due to skeletal muscle metabolism - is useful for liver function tests

Ammonia

The liver removes ammonia (which is toxic) through the circulation in what vein before it is converted to urea?

Portal Vein

Ammonia is converted to what in cases of severe liver disorders?

Glutamine

TOF: Ammonia exists as an ion in highly acidic and alkaline pH levels

False (it is an ammonium ion in PHYSIOLOGICAL pH levels)

# Analytical Methods in Determining Ammonia - uses glutamate dehydrogenase, same in urea measurement

Enzymatic Method

# Analytical Methods in Determining Ammonia - ion-exchange - ion selective electrode - spectrophotometric

Chemical

# Specimen Requirements for Ammonia - whole blood ammonia (rises/drops) rapidly following specimen collection

Rises

# Specimen Requirements for Ammonia Ammonia samples must be placed on (ice/heated blocks) to prevent the loss of ammonia

Ice

# Specimen Requirements for Ammonia Air exposure can cause ammonia to (falsely increase/decrease)

Increase

# Specimen Requirements for Ammonia What anticoagulants (2) are used to measure ammonia?

EDTA and Heparin

# Specimen Requirements for Ammonia Centrifuge the sample in what temperature range?

0-4ºC

# Specimen Requirements for Ammonia Ammonia is assayed ASAP or if frozen, it can stay stable for several days at what temperature?

-20ºC

# Specimen Requirements for Ammonia (RBCs, WBCs, or platelets) contain 2-3 times as much ammonia as plasma

RBCs

# Specimen Requirements for Ammonia Cigarette smoking is a significant contaminant and should be stopped how many hours prior to testing?

24-48 hours

# Specimen Requirements for Ammonia If sample is hemolyzed, reject immediately as this can cause a rapid (increase/decrease) of the result upon collection

Increase

# Specimen Requirements for Ammonia What is the conversion factor from ug/dL to umol/L?

0.587

Please study the images on the transes for each enzymatic/chemical reaction as these may show up on the tests, and the mere descriptions in these cards may not be enough.

Thank you.

(M) Lec 3: Non-Nitrogenous Proteins Flashcards

(140 cards)