Plasma Free Metanephrine and the Laboratory Evaluation for Pheochromocytoma

2004, Volume 15, Number 2


David F. Keren, M.D.

Pheochromocytomas are rare tumors (benign and malignant) of chromaffin cells that usually arise in the adrenal medulla, occasionally (about 10%) in paraganglia. The incidence is between 2-8/1,000,000 people. Among hypertensive patients, their prevalence is 0.13% and among individuals with incidental adrenal masses, their occurrence is 6.5%. They may occur as isolated lesions, or as part of familial endocrine syndromes. Their production of catecholamines results in hypertension that may be episodic. In addition to hypertension, clinical symptoms include sweating, headache and palpitations.

Despite their low prevalence, Pheochromocytomas need to be considered in patients with hypertension because of the potential for cure of the hypertension by excision and because of the danger of malignancy and progressive hypertension if they are missed.

Laboratory testing

Measurement of the catecholamines and their metabolic products has been performed with assays of progressively increasing sensitivity and specificity to distinguish this subset of patients from the vast majority (99.9%) with hypertension due to other causes.

For many years, spectrophotometric assays were used to measure urinary VMA and total metanephrines. These have been replaced by the more sensitive fractionated urine and serum catecholamines for detecting these tumors. In addition to their sensitivity, the HPLC methods can detect interfering substances omitting this problem with the older spectrophotometric assays. These methods are called “Fractionated” because they can distinguish epinephrine, norepinephrine, dopamine and their metanephrine metabolic products (metanephrine, normetanephrine and methoxytyramine). By detecting fractions, information is gained about possible locations of the tumors. For instance, if dopamine is increased, it increases the possibility of an extra adrenal tumor.

Despite the improvement by HPLC fractionated techniques in specificity (removes interferences), specificity and sensitivity are still not ideal. This relates to the interference by the normal production of catecholamines by sympathetic nerves and the adrenal medulla. Disease states associated with sympathoadrenal activation will cause an elevation of these values, decreasing the specificity of elevated levels determined by HPLC fractionated tests of urine or plasma.

The same cells that synthesize the catecholamines actually also perform most of the metabolism of them using monoamine oxidase to convert catecholamines to their deaminated metabolite, dihydroxyphenylglycol. But in the adrenal medulla and in pheochromocytomas, the catecholamines epinephrine and norepinephrine are converted by catechol-O-methyltransferase to the O-methylated metabolites, metanephrine and normetanephrine. By being able to detect the small amounts of these free metanephrines in plasma, one can distinguish the metabolic product that originated from an extraneuronal from one that originated from a neuronal source. Quantitatively, 90% of the metanephrine and about 40% of the normetanephrine originates in the adrenal medullary cells.

Recently there has been a significant shift from urinary fractionated metanephrines to free plasma metanephrines reflecting the increased sensitivity of plasma metanephrines in diagnosing pheochromocytoma compared to the free catecholamines. Metanephrine measurement is the total of the O-methylated metabolic product of epinephrine (metanephrine) and norepinephrine (normetanephrine). These may be detected either by HPLC with electrochemical detection or by liquid chromatography-tandem mass spectrometry (LC-MS/MS). These measurements have proven to be both a sensitive and specific method that is superior to the plasma or urinary catecholamine assays.

Why is there confusion in ordering?

Further confusing is the difference between “free” (unconjugated) metanephrine and total metanephrine (bound). When we measure urinary fractionated metanephrines or plasma total metanephrines, we split free metanephrines from their sulfate-conjugated metabolites by using acid hydrolysis. These mix with the metanephrines that were already present in a free state and they can be measured. In plasma the conjugated forms of metanephrines are about 25 times higher than the free forms, while in the urine, the conjugated forms represent about 97% of the total. Although in many cases of pheochromocytoma, the tumor cells release the intact catecholamine intermittently, free metanephrines are produced continuously, independent of the catecholamine secretion.

While one might predict that there would be a close correlation of free metanephrines with urinary metanephrines and total plasma metanephrines, there is not because of variable conjugation of sulfate to the metanephrines in the tissues of the gastrointestinal tract. Further, because of size and charge differences, free and conjugated metabolites have different renal clearance. The main source of clearance of the free metanephrines is metabolism by monoamine oxidase to 3-methoxy-4-hydroxyphenylglycol or by sulfotransferase to sulfate-conjugated metanephrines. Therefore, the free metanephrines quickly are transformed into conjugates that are only slowly eliminated by the kidneys. This slow clearance of the metabolites is a mixed blessing in the initial diagnosis of pheochromocytomas because patients with episodic pheochromocytomas will have these metabolites for longer periods of time in the plasma, but false positives will occur in patients with renal disease where the renal clearance of these normal metabolites is decreased. An important advantage of free metanephrines is that you eliminate false positives in individuals with renal impairment. In addition, the gravimetric amount helps improve specificity because in about 80% of pheochromocytomas, the excess of free plasma metanephrines is large enough to provide confidence in the diagnosis.


Plasma Free Metanephrine should be used as the screening test when pheochromocytoma is suspected. Its sensitivity is nearly 100%. When it is positive, to improve specificity, a confirmatory test with high specificity such as the fractionated 24 hour urinary metanephrines should be performed.

Potential problems with the assay

To prevent interfering endogenous synthesis of catecholamines, the patient should fast and not smoke for at least 4 hours before the specimen is drawn and the patient should drink only water during the fasting period.

Although most drugs do not directly interfere with the assay itself, they may transiently affect endogenous catecholamines, but usually, the increase of plasma free metanephrines is small. Nonetheless, to optimize the results of the plasma free metanephrine assay, the patient should discontinue epinephrine and epinephrine-like drugs at least a week before obtaining the specimen. Individuals receiving monoamine oxidase inhibitors may have an increase in their endogenous catecholamine levels. This problem will be exacerbated if the patient is also receiving a diet high in tyrosine-rich foods (bananas, cheese, etc).

Local Anesthetics, Lidocaine, Cocaine, synthetic cocaine derivatives, and anesthetic gases (halothane) may increase the levels of plasma free metanephrine.

One of the few drugs that interfere with the plasma free metanephrine assay itself is acetaminophen.

Other drugs can indirectly affect the levels of plasma free metanephrine. For instance, individuals who are undergoing withdrawal from alcohol, benzodiazepines, opiods and some central acting antihypertensive drugs (such as Clonidine) will have increased levels. Other drugs such cannabis, LSD, mescal or peyote do not generally affect the assay.

Lenders et al. recommend that the blood samples be drawn with patients in a supine position through an in-dwelling intravenous catheter.

Name of the Test:    Metanephrines, Fractionated, Free, Plasma
Reporting Nam :     Metanephrines, Fract., Free, P
Specimen Required:    At least 4mL of plasma spun down
  from an EDTA (Lavender-top) tube.
  After centrifugation, freeze 4mL of
  plasma in a plastic tube.
Reference Values:     Metanephrine Free <0.50 nmol/L
  Normetanephrine Free <0.90 nmol/L


  1. Fernandez-Calvet, L, et al.Incidence of pheochromocytoma in South Galicia, Spain . J Intern Med 1994;236:675-7.
  2. Sutton MG, et al. Prevalence of clinically unsuspected pheochromocytoma. Mayo Clin Proc 1981;56:354-60.
  3. Eisenhofer, G. et al. Plasma normetanephrine and metanephrine for detecting pheochromocytoma in von Hippel-Lindau disease and multiple endocrine neoplasia type 2. NEJM 1999;340:1872-9.
  4. Raber, W. et al. Diagnostic efficacy of unconjugated plasma metanephrines for the detection of pheochromocytoma. Arch Intern Med 2000;160:2957-63.
  5. Eisenhofer, G, et al. Plasma metanephrines are markers of pheochromocytoma produced by catechol-O-methyltransferase within tumors. J Clin Endocrin Metab 1998;83:2175-85.
  6. Pallant A, et al. Determination of plasma methoxyamines. Clin Chem Lab Med 2000;38:513-7.
  7. Eisenhofer, G. Free or total metanephrines for diagnosis of pheochromocytoma: what is the difference? (Editorial) Clin Chem 2001;47:988-9.
  8. Lagerstedt, SA, et al. Measurement of plasma free metanephrine and normetanephrine by liquid chromatography-tandem mass spectrometry for diagnosis of pheochromocytoma. Clin Chem 2004;50:603-11.
  9. Lenders JWM, et al. Biochemical diagnosis of pheochromocytoma. Which test is best? JAMA 2002;287:1427-34.
  10. Sawka AM, et al. A comparison of biochemical tests for pheochromocytoma: measurement of fractionated plasma metanephrines and catecholamines. J Clin Endocrinol Metab 2003;88:553-8.
  11. Weise M, et al. Utility of plasma free metanephrines for detecting childhood pheochromocytoma. J Clin Endocrinol Metab 2002;87:1955-60.