ND
2 SEMESTER
A.Y. 2022 - 2023
CLINICAL CHEMISTRY II
ARTERIAL BLOOD GAS ANALYSIS
ACID-BASE BALANCE
DEFINITION OF ACIDS AND BASES
 According to Arrhenius
Acid - a substance that increases concentration of
+
Hydrogen ion (H ) when dissolved in water.
Base - a substance that increases concentration of
Hydroxyl ion (OH) when dissolved in water.
Buffering system of the body
1. Bicarbonate-carbonic acid system
- Bicarbonate-carbonic acid system.
- The main buffering system of the blood which involves the
lungs and kidneys.
- Consist of carbonic acid (weak acid) and bicarbonate
(weak base)
- A 20:1 ratio of HCO3 and H2CO3 is to be maintained
 According to Bronsted and Lowry
Acid - donates proton (H+) in the reaction.
Base - accepts proton (H+) in the reaction.
 According to Lewis
Base - molecule or ion that accepts a pair of electrons to
form covalent bond.
Acid - molecule or ion that donates a pair of electrons to
form covalent bond.
2. Biphosphate-dihydrogen phosphate buffer system
3. Haemoglobin
4. Plasma proteins
pH
•
•
•
•
+
pH - potential of hydrogen ion (H ) or power of
hydrogen ion .
It determines the acidity or alkalinity of the solution
pH scale range from 0-14. 7 is neutral.
Blood pH is between 7.35-7.45 (slightly alkaline)
Clinical significance:
Acidemia – refers to decrease blood pH.
Alkalemia – refers to increase blood pH.
• Changes in blood pH are due to malfunction of either
the lungs or kidneys.
• When the lungs are impaired the kidneys compensate to
maintain blood pH. (metabolic compensation)
• When kidneys are impaired the lungs compensate to
maintain blood pH. (respiratory compensation).
Hydrogen ion concentration and pH
Inverse relationship between hydrogen ion and pH
Types of ions (based electric field migration)
1. Cations – (+) charge ions moved
toward the cathode (negative
side)
2. Anions – (-) charge ions
moved toward the anode
(positive side)
General functions of electrolytes
•
•
•
•
•
•
•
Consist of : Weak acids
Conjugate base
-
LABORATORY METHOD OF DETERMINATION
-
These are substance that resists changes in blood pH.
Capable of neutralizing acids or bases to maintain the
original pH of a solution.
Present in all body fluids, ICF and ECF.
+
Distribution in the body
Body fluids are mainly
composed of water located in
different compartments.
A. Intracellular fluid (ICF) – 2/3
of the total body water.
Located inside the cells.
B. Extracellular fluid (ECF) – 1/3
of the Total body weight.
1. Intravascular fluid – plasma
2. Interstitial fluid – surrounds the cells and tissues.
BUFFERING SYSTEM OF THE BODY
Buffers
•
•
-
Acid base balance – ( HCO3 , K , Cl )
+ +
Osmotic pressure regulation and body hydration- ( Na , K , Cl)
2+
2+
2+
Enzyme co-factors and blood coagulation-( Mg , Ca , Zn )
Neuromuscle excitability and Heart muscle rhythm and
2+
2+ +
contractility (Mg , Ca , K ).
Buffering system of the blood- ( HCO3 )
2+
ATP production from glucose- ( Mg , PO4 )
Electrolytes can be measured by the following:
1. Colorimetric method
2. Flame emission photometry
3. Atomic absorption photometry
4. Potentiometry - Ion-selective electrode (ISE)
5. Gravimetric
6. Turbidimetric
LEIZA MORIE
1
ND
2 SEMESTER
A.Y. 2022 - 2023
+
CLINICAL CHEMISTRY II
ARTERIAL BLOOD GAS ANALYSIS
Sodium (Na )
Most abundant cation (90 %) in ECF less concentration in ICF.
Regulated by hormone aldosterone through reabsorption from
the kidneys.
Functions:
a. Maintain osmotic pressure, this prevents the cell from
bursting.
b. Water balance and maintain blood pressure.
c. Neuromuscular excitability
Levels are regulated by:

ADH – water retention causes decrease of Na (dilutional
hyponatremia).

Aldosterone – increase secretion; increase Na reapsortion
(hypernatremia)

Thirst
Normal value of sodium
1. Serum – 135- 148 mEq/L
2. CSF – 138-155 mEq/L
Clinical significance
 Hyponatremia (< 135 mEq/L)
- Due to increase sodium loss or increase water retention.
- One of the most common electrolyte disorders.
Symptoms: nausea, vomiting, muscle weakness, lethargy, headache, and
ataxia.
- <125 mEq/L – severe neuropsychiatric symptoms.
Causes: Dehydration, vomiting, diarrhea, severe burns, metabolic acidosis,
renal disease, Addison’s disease, syndrome of inappropriate ADH secretion
(SIADH), nephrotic syndrome
 Pseudohyponatremia
- Marked hemolysis decreased sodium level due to dilutional
effect.
- Hyperlipidemia and hyperprotenemia may also falsely
decrease serum sodium (due dilutional effect).
 Hypernatremia (> 145 mEq/L)
Increase sodium concentration due to loss of water, gain of
sodium or both.
Less common.
Symptoms: altered mental status, lethargy, irritability, seizures, muscle
twitching, hyper reflexes, difficulty in breathing and increase thirst.
Causes: diabetes insipidus, renal disease, decrease water intake, fever,
vomiting, alkalosis, severe burns, hyperadrenalism, diabetic coma.
Specimen of choice:
a. Serum
b. Plasma
c. 24 urine
d. CSF
e. Sweat
Anticoagulant: Heparin
Specimen consideration
1. Avoid hemolysis
2. Used for pure water to avoid trace contamination
3. Smoking increases sodium level
4. Prolong standing increases sodium level (shift to intracellular
fluid)
5. oxalate decreases Na level
6. exercise decreases Na due to sweat
7. thumb or fingers must not use to cover tubes because it
contains sodium
Laboratory method
Colorimetric method (Albanese and Lein)
1. sodium + zinc uranyl acetate = sodium uranyl acetate precipitate
2. sodium uranyl acetate precipitate + water + sodium salicylate =
yellow precipitate
Flame emission photometry – produce yellow color when exposed to
flame.
Ions selective electrode (ISE) – used glass aluminum silicate
Reference value
 serum: 135-145 mmol/L
 24 H urine: 40-220 mmol/L
 CSF: 138-150 mmol/L
Conversion factor: mmol/L to mEq/L= 1.0
+
Potassium (K )
Major cation in ICF and less in ECF.
No renal threshold
Functions
1. Muscle contraction both skeletal and cardiac.
2. Respiration.
3. Muscle irritability
Reference value
 Serum: 3.5-5.5 mmol/L (adult) Infant: 4.0 – 5.9 mEq/L
 24 H urine: 25-125 mmol/L
mmol/L- mEq/L= 1.0
- Increase or decrease level of K may result to muscle weakness,
paralysis and cessation of cardiac contraction.
Clinical significance
+
 Hypokalemia – decrease level of K
Symptoms: weakness, fatigue, constipation, paralysis, difficulty in
breathing.
Causes: diarrhea, vomiting, renal disease, tumor, malabsorption, Cushing
syndrome, hypomagnesemia, hyperaldosteronism, alkalosis.
 3.0-3.4- mild hypokalemia
The most common cause of hypokalemia due to increase activity of
aldosterone (hyperaldosteronism)
Diarrhea – causes direct K loss in the stool.
Vomiting – mainly result of K loss in the urine.
-
Effects in cardiac muscle
1. Decreases cell excitability resulting to arrhythmia and
paralysis
2. The heart may experience cessation in contraction in either
hyperkalemia or hypokalemia.

Alkalosis: decrease serum potassium
Pseudohypokalemia
Leukocytosis cause falsely decreases K. as seen in leukemia if
sample left at room temperature.
Causes: Hemolysis, thrombocytosis, prolong tourniquet
application, fist clenching, blood stored in ice, IV fluid.
+
 Hyperkalemia – Increased level of K
Symptoms: muscle weakness, tingling, numbness, mental confusion,
cardiac arrhythmias.
Causes: renal disease, Addison’s disease, diuretics, muscle injury, acidosis,
hypo aldosteronism, haemolytic disease.
The most common cause of hyperkalemia is impaired renal
excretion such as aldosterone deficiency and renal failure.
Hyperkalemia
LEIZA MORIE
2
ND
2 SEMESTER
A.Y. 2022 - 2023
CLINICAL CHEMISTRY II
ARTERIAL BLOOD GAS ANALYSIS
1.
2.
3.
Acidosis : potassium increases
Low insulin causes elevated serum potassium
Hyperkalemic drugs: captopril, digoxin, heparin therapy.
Effects on cardiac muscle
1. 8 mmol/L – lack of muscle excitability (ability to respond to
stimulus)
2. 10 mmol/L – can lead to cardiac arrest.
Specimen consideration
1. Hemolyzed sample must be avoided to prevent falsely elevated
K.
2. ( K level inside the cell is very high)
3. Exercise increases K level to avoid this, let the patient rest for
a few minutes before drawing blood sample.
4. Used pure water in analysis.
5. K level is slightly lower in plasma than in serum
Laboratory method
Colorimetric method (Lockhead and Purcell) – turbidity is proportional to
the K concentration.
Flame emission spectrophotometry – produces violet color when expose
to flame.
Ion-selective electrode (ISE) – used valinomycin gel
Atomic absorption spectrophotometry
Causes: hypothyroidism, hyperaldosteronism, bone cancer, antacids,
dehydration, Addison’s disease, acute renal failure.
Specimen:
a.
b.
c.
d.
e.
Specimen consideration
1. No venous stasis or tourniquet during collection
2. Avoid hemolysis because RBC contains 10x magnesium than
serum/plasma
3. Physiologic increase in women with ecclampsia.
Laboratory method
Colorimetric method
1. Calmagite – produce violet color
2. Methyl – thymol blue
3. Titan yellow (Dye-lake method)
4. formazan dye (vitros machine)
ISE
AAS – reference method
Fluorometric analysis – fluorescent compound
+
Magnesium (Mg )
2 most abundant cation in the ICF.
53%- found in the bone, 46 %- found in muscle and other organs.
Supplied by diet.
3 form in the blood
a. 55 %- free magnesium
b. 30 %- protein-bound
c. 15 % - complexed Mg
Functions
1. Muscle contraction and heart rhythm.
2. Enzyme co factors ( glycolysis, synthesis of carbohydrates,
lipids, proteins etc)
3. Maintains the structure of DNA, RNA and ribosomes.
nd
Factors affecting magnesium in the blood

PTH

Increases renal reabsorption and intestinal absorption.

Aldosterone and thyroxine

Increases renal excretion.
Reference value
 0.63- 1.0 mmol/L
 5.0 mmol/L- life threatening
Clinical significance
 Hypomagnesemia
most commonly observed in ICU patients and undergoing diuretic
therapy.
Symptoms: arrhythmia, psychosis, weakness, cramps, depression, seizure,
tetany, paralysis.
Causes: malabsorption syndrome (sprue), malnutrition, chronic alcoholism,
hyperaldosteronism, hyperthyroidism, acute renal failure, severe
diarrhea.
 Hypermagnesemia
Symptoms: hypotension, bradycardia, increase temperature, nausea,
vomiting, lethargy.
Serum
Plasma ( lithium heparinized)
24 urine
Avoid using EDTA, oxalate, and citrate.
Avoid hemolysis.
2+
-
Calcium (Ca )
Only 1 % found in the blood the rest in the bones. Most abundant
cation in the body
Blood:
1.
free calcium (ionized) the active form- 50 %
2. complex Ca-10 %
3. protein bound – 40 %
- Calcium is absorbed in the small intestine.
- Regulators of blood Cacium: Parathyroid hormone, calcitonin, and
Vitamin D.
- Renal threshold: >10 mg/dl
Functions
1.
2.
3.
4.
5.
Neurotransmitter regulator.
Helps for proper contraction of the heart.
For blood coagulation.
Enzyme activator.
Maintain bone and teeth structure.
Factors influencing serum calcium level
 PTH (Parathyroid hormone)
- mobilizing calcium from the bones
- increase synthesis of 1,25-dihydroxycholecalciferol
 Calcitonin
- produced by p-follicular cells of the thyroid gland
- inhibits bone resorption by regulating the activity of
osteoclasts
- antagonist to PTH
 Active Vitamin D3 (active calciferol)
- undergoes
hydroxylation
producing
1,25dihydroxycholecalciferol
 Plasma protein
- direct relationship with calcium
 Serum/Plasma Phosphate
- inverse relationship with calcium
Other factors affecting calcium level
LEIZA MORIE
3
ND
2 SEMESTER
A.Y. 2022 - 2023
CLINICAL CHEMISTRY II
ARTERIAL BLOOD GAS ANALYSIS
Acidosis: decreases protein binding resulting to increase calcium
Alkalosis: increases protein binding resulting to decrease calcium.

Hypoclorenemia
Clinical Significance
Causes: DKA, aldosterone deficiency, vomiting, high serum bicarbonate.
Metabolic alkalosis, diuretics.


Hypocalcemia
Symptoms: muscle cramps, tetany, seizures, and arrhythmia.
Causes: primary hypothyroidism, vit D deficiency, renal failure, Fanconi’s
syndrome, hypoalbuminemia, pacreatitis, alkalosis

Hypercalcemia
Symptoms: often asymptomatic, severe cases include mild weakness,
lethargy, GI symptoms and renal discomfort.
Causes: hyperthyroidism, acidosis, lung cancer, increase vit D, multiple
myeloma, sarcoidosis.
Specimen
1. Serum
2. Plasma- lithium anticoagulant
3. 24-hour urine
Reference value
Total calcium
2.20-2.70 mmol/L (children)
2.15-2.50 mmol/L (adult)
Ionized calcium
1.20-1.38 mmol/L (children)
1.16-1.32 mmol/L (adult)
Urine
2.50-7.50 mmol/day
-
Phosphate (PO4)
Intracellular anion.
most are in the form of inorganic phosphate (serum)
In complex with magnesium and calcium
70% of plasma inorganic phosphorus is organic and 30% are
inorganic.
Majority of inorganic phosphate is excreted in urine and PTH
favors urinary loss of phosphorus.
Absorption in small intestines.
Functions:
1. For intermediary metabolism of carbohydrates
2. Components of phospholipids, nucleic acid, nucleotide and
inorganic phosphate esters
3. Bone mineralization
-
Functions:
1. Maintain water balance and osmotic pressure.
2. Acid-base balance
3. Maintenance of electrolyte balance/electroneutrality
(Chloride shift)-exchange with HCO3 across the RBC membrane.
4. Hydration.
Clinical Significance
Serum
Plasma- lithium heparinized
Urine
CSF
Sweat
Tears
Colorimetric method
1. Schales and schales (mercuric nitrate titration method) produces paint violet color.
2. Mercuric thiocynate and ferric nitatre produce reddish color
ISE method – used silver chloride- silver sulfide membrane.
Coulometric (amphometric method) – measure by Chloridometer with
silver ion
-
Chloride (Cl )
Major anion in the ECF.
Counter ion of Na- always accompany Na to maintain electric
neutrality.
Lost or excreted in sweat.
Only anion that enzyme activator – activates amylase
Regulated by aldosterone.
Reference range
Plasma: 98- 107 mmol/L
24 H urine: 110- 250 mmol/dl
Specimen:
a.
b.
c.
d.
e.
f.
-
ANIONS
-
Causes: metabolic acidosis, renal tubular acidosis, excess loss of
bicarbonate, cystic fibrosis, CHF, hyperthyroidoism, severe diarrhea.
Laboratory determination
Laboratory determination
1. Orthocresolphthalein complexone dyes (colorimetric)
2. AAS- reference method
3. ISE- ion exchanger membrane or ionophore
4. Clark collip precipitation method- measure oxalic acid as the
end point
5. Ferro Ham chloroanilic acid precipitation method
6. Compleximetric- removal of calcium by EDTA
-
Hyperchlorenemia
Clinical Significance
 Hypophospathemia
Causes:
a. hyperparathyroidism
b. rickets
c.
osteomalacia
d. Fanconi’s syndrome
 Hyperphosphatemia
Causes:
a. hypoparathyroidism
b. increase Vit D
c. hypersecretion of growth hormone
d. uremia
e. glumerulonephritis
Reference range
At Birth
Children
Adult
1.34 – 3.35 mmol/L
1.28 – 2.24 mmol/L
0.96 – 1.44 mmol/L
Specimen
LEIZA MORIE
4
ND
2 SEMESTER
A.Y. 2022 - 2023
CLINICAL CHEMISTRY II
ARTERIAL BLOOD GAS ANALYSIS
1.
2.
3.
4.
5.
Serum
Plasma-lithium heparinized
24-hour urine
Avoid hemolysis
Circadian variation- highest in the morning
Laboratory determination
FISKE-SUBARROW METHOD (spectrophotometry)
Reagent: Molybdate
End color: Blue
-
Bicarbonate (HCO3 )
2 most abundant anion in ECF
nd
-
Functions:
Major buffering system of the blood.
Exchange with Cl across the RBC to maintain ionic charge
neutrality within the cell (chloride shift).
Clinical Significance
Decrease in:
Metabolic acidosis
+
HCO3 combines with hydrogen ion (H ) to produce CO2 which is
exhaled by the lungs.
SUMMARY
ELECTROLYTES
Anions
(-) charge
Cations
(+) charge
+
+
2+
-
2+
-
2-
Cl , HCO , PO
Na , K , Mg , Ca
3
4
NORMAL VALUES
Note: Normal values may vary slightly from other laboratories
Electrolytes
+
Na
N. V.
135-145 mEq/L
Cl
-
98-107 mEq/L
+
3.4-5.0 mEq/L
K
2+
Ca
Total- 2.15-2.50 mmol/L
Ionized- 1.16-1.32 mmol/L
Mg
2+
0.63- 1.0 mmol/L
PO4
2-
0.87- 1.45 mmol/L
HCO3
-
22-26 mEq/L
LEIZA MORIE
5