Estimation of calcium and phosphorus

DUDEJA

Three hormones, PTH, vitamin D, and calcitonin, are known to regulate serum Ca2+ by altering their secretion rate in response to changes in ionized Ca2+

Distribution

About 99% of Ca2+ in the body is part of bone. The remaining 1% is mostly in the blood and other ECF. Little is in the cytosol of most cells.

Ca2+ in blood is distributed among several forms. About 45% circulates as free Ca2+ ions (referred to as ionized Ca2+), 40% is bound to protein, mostly albumin, and 15% is bound to anions, such as HCO3 −, citrate, HCO3 −, and lactate

Symptoms of hypocalcemia Neuromuscular irritability and cardiac irregularities are the primary

groups of symptoms that occur with hypocalcemia.Neuromuscular symptoms include paresthesia, muscle cramps, tetany,and seizures.

Cardiac symptoms may include arrhythmiaor heart block. Symptoms usually occur with severehypocalcemia, in which total Ca2+ levels are below 1.88mmol/L (7.5 mg/dL).

Symptoms of hypercalcemiaModerate or severe Ca2+ elevations include NEUROLOGIC, GI, AND

RENAL SYMPTOMS. Neurologic symptoms may include mild drowsiness or weakness,

depression, lethargy, and coma. GI symptoms may include constipation, nausea, vomiting, anorexia,

and peptic ulcer disease. Hypercalcemia may cause renal symptoms of nephrolithiasis and

nephrocalcinosis. Hypercalciuria can result in nephrogenic diabetes insipidus, which

causes polyuria that results in hypovolemia, which further aggravates the hypercalcemia.

Hypercalcemia can also cause symptoms of digitalis toxicity.

MethodsThe two commonly used methods for total Ca2+ analysis use

either ortho-cresolphthalein complexone (CPC) or arsenazo III dye to form a complex with Ca2+. Prior to the dye-binding reaction, Ca2+ is released from its protein carrier and complexes by acidification of the sample. The CPC method uses 8-hydroxyquinoline to prevent Mg2+ interference.

AAS remains the reference method for total Ca2.

Current commercial analyzers that measure ionized/ free Ca2+ use ISEs for this measurement. These systems may use membranes impregnated with special molecules that selectively, but reversibly, bind Ca2+ ions. As Ca2+ binds to these membranes, an electric potential develops across the membrane that is proportional to the ionized Ca2+ concentration.

PHOSPHATE

An adult has about 600 g or approximately 20 mol of phosphorus in inorganic and organic phosphates, of which about 85% is in the skeleton, and the rest is principally in soft tissue.

Plasma contains both inorganic and organic phosphate, but only inorganic phosphate is measured. Inorganic phosphate exists as both monovalent (H2PO4 −) and divalent (HPO4 2−) phosphate anions.

Approximately 10% of the phosphate in serum is protein-bound; 35% is complexed with sodium, calcium, and magnesium; and the remainder, or 55%, is free.

In the soft tissue, most phosphate is cellular. Although both inorganic and organic phosphate is present in cells, most is organic and is incorporated into nucleic acids, phospholipids, phosphoproteins, and high-energy compounds involved in metabolism like ATP.

Method of demonstration

Most methods used to measure serum inorganic phosphate are based on the reaction of phosphate ions with ammonium molybdate to form a phosphomolybdate complex that is then measured by a spectrophotometer

The colorless phosphomolybdate complex may be measured directly by ultraviolet absorption (340 nm) or reduced to molybdenum blue and measured at 600 to 700 nm.

An acidic pH is necessary for the formation of complexes, but it must be controlled because both complex formation and reduction of molybdate are dependent on pH. A less acidic pH can result in spontaneous reduction of molybdate.

Phosphate concentrations can also be determined by several other procedures, including the vanadate-molybdate and enzymatic methods.

Vanadate and molybdate form a yellow complex with phosphate at acid pH, but the method tends to overestimate inorganic phosphate because of hydrolysis of organic esters.

Interferences

Depending on the method used, positive or negative interference has been noted with hemolyzed,

icteric, & lipemic specimens. Mannitol, fluoride, and monoclonal

immunoglobulins have also been reported to interfere. Glassware should be properly cleaned and rinsed

because phosphate is a common component of many detergents