Serum calcium. Routine serum calcium assay measures the total serum calcium value. Total serum calcium contains about 50% bound calcium (literature range, 35%-55%) and about 50% nonbound calcium (literature range, 35%-65%). (Traditionally, nonbound calcium was called “ionized” calcium and is also known as “free” or “dialyzable” calcium.) Bound calcium is subdivided into calcium bound to protein and calcium complexed to nonprotein compounds. About 45% of total calcium (30%-50%) is protein-bound, of which 70%-80% is bound to albumin. The remaining 5% (5%-15%) of total calcium is complexed to ions such as citrate, phosphate, sulfate, and bicarbonate, which are not part of the serum proteins. Ionized calcium levels can be measured directly by ion-selective electrode techniques or less accurately can be estimated from total serum calcium and albumin or total protein values using certain formulas. The most commonly used calcium correction formula is that of R.B. Payne:

Adjusted calcium = (measured calcium – serum albumin) + 4.0

with calcium in mg/100 ml and albumin in g/100 ml. In international system (SI) units, the formula reads:

Adjusted calcium = (calcium – 0.025 albumin) + 1.0

with calcium in mmol/L and albumin in g/L. Ionized calcium values are affected by serum pH (a decreased of 0.1 pH unit increases ionization by 1.5%-2.5%). If serum is exposed to air and stands too long, the pH slowly increases. There is a small diurnal variation in ionized calcium, with the peak most often about 9 P.M. and the nadir about 9 A.M. There also is a small diurnal variation in urine calcium, with the peak most often about 11 P.M. and the nadir about 11 A.M.

Ionized calcium is not affected by changes in serum albumin concentration, which is a significant advantage over total calcium assay. A decrease in the serum albumin level by 1 gm/100 ml produces an approximate decrease in the (total) serum calcium level of approximately 0.8 mg/100 ml from previous levels (this is an average value, and could be more or less in any individual patient). Since a decrease in the serum albumin level is frequent in patients with severe acute or chronic illness, an artifactual decrease of the serum calcium level is likewise frequent in hospitalized patients. The ionized calcium value is regarded by many investigators as more sensitive and reliable than the total calcium value in detection of PHPT. A certain number of their PHPT patients had elevated ionized calcium levels but normal total calcium levels. However, some investigators do not find that ionized calcium materially assists detection or diagnosis of PHPT. Also, certain conditions that produce hypercalcemia, such as myeloma, sarcoidosis, hypervitaminosis D, and metastatic carcinoma to bone, may in some cases be associated with increased ionized calcium levels. Most laboratories do not have the equipment necessary to perform ionized calcium assay, and although there are formulas that estimate ionized calcium from total calcium plus serum protein levels, there is disagreement in the literature concerning which formula is best. There is also disagreement whether such estimates are reliable enough to be used in the diagnosis of PHPT. The consensus in the literature seems to be that ionized calcium may be helpful in the diagnosis of PHPT in a minority of patients, such as those with borderline total calcium values or those with hypoalbuminemia, and is best determined using ion-selective electrode methodology.

Serum phosphate. Decreased serum phosphate is one of the classic biochemical findings in PHPT. Phosphate usually is measured as the phosphorus ion. Only about 10%-15% is protein bound. There is a diurnal rhythm, with higher values in the afternoon and evening, which may be as much as double those in the morning. Serum phosphate (phosphorus) has not proved as useful as the earliest studies suggested, since phosphate levels in PHPT that are below the population reference range tend to be limited to patients with more severe disease. In fact, the serum phosphate level is decreased in only about 40%-50% of PHPT cases (literature range, 22%-80%). The reference range is fairly wide, which can mask small decreases; and some conditions such as a high-phosphate or a low-calcium diet increase serum phosphate levels. Renal dysfunction severe enough to produce an elevated blood urea nitrogen (BUN) level raises the serum phosphate level. Various conditions other than PHPT can decrease serum phosphate levels (discussed later in this chapter). Among those associated with hypercalcemia and hypophosphatemia, besides PHPT, are some patients with malignancy and occasional patients with sarcoidosis, myeloma, hyperthyroidism, and vitamin D intoxication.

Serum alkaline phosphatase. Alkaline phosphatase in nonneoplastic calcium disorders is an index of bone involvement. X-ray bone abnormalities in PHPT are reported in 23%-36% of patients, with most of the relatively few reports being in the older literature. X-ray studies of the fingers demonstrate the most typical changes. It is estimated that alkaline phosphatase elevation occurs in about 95% of patients with PHPT who have bone x-ray changes but in only about 10%-15% of those who do not (therefore, there would be an ALP level increase in 20% to 30% of all PHPT cases; however, some find x-ray changes in present-day PHPT in only 15% of cases). Metastatic malignancy can produce elevated alkaline phosphatase levels due to either bone or liver involvement.

Urine calcium excretion. Excretion of calcium in urine is increased in about 75% of patients with PHPT (literature range, 50%-91%). In addition to PHPT, a considerable number of other conditions may produce hypercalciuria (e.g., idiopathic hypercalciuria, said to be present in nearly 5% of the population; bone immobilization syndrome; Cushing’s syndrome; milk-alkali syndrome; hypervitaminosis D; renal tubular acidosis; and sarcoidosis).

Assay is performed on 24-hour urine specimens. There is disagreement in the literature on whether to collect the specimens with the patient on a normal diet, a normal diet minus milk, cheese, and other milk products, or a standard 200-mg low calcium diet. Most investigators and urologists seem to prefer a normal diet, at least for screening purposes. Reference ranges differ according to type of diet. The Sulkowitch test is a semiquantitative chemical procedure for urine calcium measurement that was widely used before 1960 but is rarely performed today.

Urine phosphate excretion. The older literature states that phosphate excretion is increased in most patients with PHPT. There are surprisingly little data on this subject in recent literature. However, hyperphosphaturia probably is not as frequent today, just as decreased serum phosphate levels are seen much less frequently. Increased urine phosphate excretion in PHPT is expected in 70%-75% of cases. Phosphate depletion (due to prolonged vomiting, nasogastric suction, or ingestion of aluminum-type antacids) and chronic renal disease can reduce or eliminate phosphate hyperexcretion. Conditions that can produce increased urine phosphate excretion besides PHPT include renal triple phosphate lithiasis, osteomalacia, and, in some patients, hyperthyroidism, sarcoidosis, Cushing’s syndrome, and malignancy. Reference values depend on diet.