Chloride, Bicarbonate, Calcium, Magnesium, and Phosphate

Four electrolytes complete the core panel after sodium and potassium. Chloride is the principal extracellular anion and travels with bicarbonate through exchange reactions that preserve electroneutrality. Total CO₂ on a chemistry panel is, at physiologic pH, effectively a bicarbonate measurement. Calcium, magnesium, and phosphate share regulation through parathyroid hormone and vitamin D, share chelation restrictions on specimen tubes, and share the property that their total serum measurements can misrepresent the active fraction in acute illness. Each result is interpreted against the reporting laboratory’s verified interval.¹

Chloride

Chloride is the principal extracellular anion. It accompanies sodium or exchanges with bicarbonate to preserve electroneutrality, osmolality, and volume. Proximal sodium reabsorption is chloride-limited, and the chloride shift exchanges intracellular bicarbonate for plasma chloride as erythrocytes load CO₂. Clinically, chloride generally mirrors sodium, but hyperchloremia accompanies bicarbonate loss in diarrhea, renal tubular acidosis, and the other normal-anion-gap acidoses. Hypochloremia follows prolonged vomiting, diabetic ketoacidosis, aldosterone deficiency, salt-losing nephropathy, or expanded-bicarbonate states such as compensated respiratory acidosis and metabolic alkalosis.¹

Specimen and method. Serum or lithium-heparin plasma; hemolysis matters little except grossly. The routine method is ISE potentiometry with an ion-exchange membrane. Amperometric-coulometric and mercurimetric titration persist in niche use.

Total CO₂, the chemistry-panel bicarbonate

Venous serum or plasma total CO₂ is the second-most-abundant extracellular anion and the principal blood buffer. The total CO₂ measurement includes bicarbonate, dissolved CO₂, and carbonic acid, of which more than 90% is bicarbonate at physiologic pH, so the panel value effectively is bicarbonate. Proximal tubules reclaim about 85% of filtered bicarbonate through the hydrogen-ion-secretion and carbonic-anhydrase mechanism developed in the acid-base module. Alkalosis promotes urinary bicarbonate excretion and acidosis maximizes reclamation. Uncapped specimens lose dissolved CO₂ at roughly 6 mmol/L per hour, so prompt, capped analysis is required.¹

Methods. Two approaches exist. The electrode method acidifies the specimen, converting all CO₂ species to gas measured by a pCO₂ electrode. The enzymatic method alkalinizes instead, driving everything to bicarbonate:

Phosphoenolpyruvate + HCO₃^- → oxaloacetate + PO₄^3- (PEP carboxylase)

Oxaloacetate + NADH + H⁺ → malate + NAD⁺ (malate dehydrogenase)

Falling 340-nm NADH absorbance is proportional to bicarbonate. The venous serum and plasma total CO₂ reference interval is 22 to 28 mmol/L in the source teaching set; each laboratory reports against its own verified interval.

Examination reference ranges. The ASCP BOC certification examination uses a separate composite range set: sodium 136–145 mmol/L, total CO₂ 22–33 mmol/L, arterial pCO₂ 35–44 mm Hg, arterial pO₂ above 80 mm Hg, and arterial bicarbonate 23–29 mmol/L (June 9, 2026 guideline). Answer examination items from these values and report patient results from the laboratory’s own verified reference interval. The complete composite set is in the examination reference ranges topic.²

Calcium

About 99% of body calcium, roughly 1 kg, is skeletal. Blood calcium splits three ways: about 45% free ionized Ca²⁺, the physiologically active fraction; about 40% albumin-bound; and about 15% complexed with bicarbonate, citrate, or lactate. Ionized calcium averages about 1.18 mmol/L; its depression impairs myocardial contractility and can progress to tetany.¹

Because albumin, citrate, bicarbonate, and lactate all swing during surgery and critical illness, total calcium may misrepresent calcium status in those settings, and ionized calcium is the perioperative and ICU measurement of choice. Ionized calcium cannot be reliably back-derived from total calcium in acute illness. In stable outpatients, a correction may improve interpretability:¹

Corrected total Ca (mg/dL) = measured total Ca + 0.8 × (4.0 − albumin [g/dL])

Worked example. Total calcium is 7.9 mg/dL with albumin 2.5 g/dL. Corrected calcium = 7.9 + 0.8 × (4.0 − 2.5) = 7.9 + 1.2 = 9.1 mg/dL. The calculation illustrates how low albumin can depress total calcium even when the estimated albumin-adjusted value falls within the expected range; the correction remains an estimate that assumes otherwise normal binding conditions.

Regulation. Three hormones act on gut, kidney, and bone. Parathyroid hormone (PTH), released as ionized calcium falls, drives osteoclastic resorption, renal tubular Ca²⁺ reabsorption, renal phosphate excretion, and renal 1α-hydroxylase activation. 1,25-dihydroxyvitamin D₃, produced by hepatic and renal hydroxylation of dietary or cutaneous vitamin D₃, drives intestinal calcium and phosphate absorption and cooperates with PTH on bone. Calcitonin, from thyroid C cells, opposes both, lowering calcium, though its minute-to-minute physiologic role is minor.

Hypocalcemia and hypercalcemia

Perioperative and ICU monitoring favors ionized calcium because citrated blood products, bicarbonate infusions, and acute protein shifts render calculated corrections unreliable. Neonatal ionized calcium peaks at birth, falls 10 to 20% over the first 1 to 3 days from immature PTH and vitamin-D axes plus transient hyperphosphatemia and hypomagnesemia, then stabilizes slightly above adult levels within about a week.¹

Determination. Total calcium uses serum or lithium-heparin plasma collected without prolonged stasis. EDTA and oxalate are prohibited because they chelate calcium. Ionized calcium requires anaerobic handling, since CO₂ loss raises pH and shifts calcium onto albumin, falsely lowering the ionized fraction; therefore use anaerobically sealed heparinized whole blood, or serum from tubes clotted and spun within 30 minutes. Liquid heparin itself binds calcium (25 IU/mL lowers ionized Ca²⁺ about 3%), so balanced dry-heparin tubes are used. Total calcium is measured by colorimetry, using o-cresolphthalein complexone shielded from Mg²⁺ by 8-hydroxyquinoline or arsenazo III, with atomic absorption spectrophotometry as the reference method. Ionized calcium is measured exclusively by calcium-selective ISE.

Magnesium

Mg²⁺ is the fourth most abundant body cation and second most abundant intracellularly, behind potassium. A 70-kg adult holds about 1 mol (24 g), roughly 53% in bone, 46% in muscle and soft tissue, and under 1% in serum and erythrocytes. Of serum Mg²⁺, about one-third is protein-bound, mostly to albumin, about 60% is free and active, and about 5% is complexed. Mg²⁺ is a cofactor for more than 300 enzymes spanning glycolysis, ion pumping, neuromuscular transmission, and macromolecular synthesis.¹

Regulation. Intestinal absorption runs 20 to 65% of intake according to need. Renally, only 25 to 30% of non-protein-bound filtered Mg²⁺ is reclaimed proximally, unlike sodium’s 60 to 75%; the thick ascending limb of Henle reclaims 50 to 60%, the principal regulatory site, with 2 to 5% more distally. The renal threshold, 0.60 to 0.85 mmol/L, sits near normal serum levels, so a slight excess spills immediately into urine; normally only 4 to 6% of the filtered load reaches urine. PTH promotes both renal and intestinal Mg²⁺ handling.

Hypomagnesemia and hypermagnesemia

Hypomagnesemia is usually silent above 0.5 mmol/L. Below that concentration, manifestations may be cardiovascular (arrhythmia, hypertension, potentiated digitalis toxicity), neuromuscular (tremor, tetany, seizure, paralysis), psychiatric (agitation, psychosis), or metabolic. Metabolic findings include hypokalemia in roughly 40% of hypokalemic inpatients, plus hypocalcemia, hypophosphatemia, and hyponatremia. These companion deficits persist until magnesium is repleted. Hypermagnesemia above about 1.5 mmol/L causes hypotension, bradycardia, flushing, and nausea. Above about 5.0 mmol/L, it causes ECG changes, heart block, respiratory paralysis, and loss of deep-tendon reflexes. It also suppresses PTH, inviting hypocalcemia, and impairs hemostasis by competing with calcium in thrombin generation and platelet adhesion.¹

Determination. Use nonhemolyzed serum or lithium-heparin plasma, promptly separated, because erythrocytes concentrate Mg²⁺ roughly tenfold over plasma. Oxalate, citrate, and EDTA chelate and are excluded. Routine methods are colorimetric dye binding with calmagite, formazan, or methylthymol blue plus Ca²⁺-masking agents; atomic absorption spectrophotometry is the reference method. Total serum magnesium can appear normal while tissue stores run about 20% depleted, because only a minority of body Mg²⁺ is extracellular.

Phosphate

Inorganic phosphate forms part of nucleic acids, coenzymes, and the high-energy phosphates ATP, creatine phosphate, and phosphoenolpyruvate. Deficiency reduces cellular ATP and also lowers red-cell 2,3-DPG, left-shifting the oxyhemoglobin curve and impairing O₂ delivery. Roughly 80% of body phosphate is skeletal; intracellularly it is the dominant anion.¹

Regulation. PTH lowers blood phosphate by increasing renal excretion, the dominant control. Vitamin D raises it through intestinal absorption and renal reabsorption. Growth hormone decreases renal excretion, so GH excess produces hyperphosphatemia. Transcellular shifts, where phosphorylation reactions pull phosphate into cells, complicate acute interpretation.

Hypophosphatemia and hyperphosphatemia

Hypophosphatemia affects 1 to 5% of hospitalized patients, rising to 20 to 40% in DKA, COPD, asthma, malignancy, prolonged parenteral nutrition, inflammatory bowel disease, anorexia nervosa, and alcoholism, and to 60 to 80% in septic ICU patients. Mechanisms are increased renal excretion, as in hyperparathyroidism, and decreased absorption, as in vitamin D deficiency or phosphate-binding antacids. Severe depletion, below 0.3 mmol/L, carries substantially higher mortality than mild-to-moderate cases. Hyperphosphatemia is chiefly a renal-failure disease where excretion fails; other contributors are excess intake, as in infants on cow’s-milk formula; cellular release in severe infection, intense exercise, tumor lysis, hemolysis, or lymphoblastic leukemia, where immature lymphoblasts carry about four times the phosphate of mature lymphocytes; and hypoparathyroidism, which removes PTH-driven excretion.¹

Determination. Use serum or lithium-heparin plasma. Oxalate, citrate, and EDTA interfere and are barred. Hemolysis falsely elevates the result because red-cell phosphate is abundant. Serum phosphate has diurnal variation, generally reaching its nadir in the morning and rising later with a nocturnal peak, so collection timing should be standardized; urine phosphorus requires a 24-hour collection. The method measures the ammonium phosphomolybdate complex directly by UV absorbance at 340 nm or after reduction to molybdenum blue at 600 to 700 nm.

References

1. Rifai N, Chiu RWK, Young I, Burnham CAD, Wittwer CT, eds. Tietz Textbook of Laboratory Medicine. 7th ed. Elsevier; 2023.
2. American Society for Clinical Pathology Board of Certification. MLS(ASCP) and MLS(ASCP)ⁱ Content Guideline. Revised June 9, 2026. Examination composite reference ranges, pp. 16-17.