Metanephrine, catecholamine

Chan ECY, Wee PY, Ho PY, Ho PC. 2000. High-performance liquid chromatographic assay for catecholamines and metanephrines using fluorimetric detection with pre-column 9-fl.uorenylmethyloxycarbonyl chloride derivatiza-tion. J Chromatogr B 749 179... 

Several additional useful publications demonstrating practical applications of CE/MS methods for neurotransmitter analysis and neuropharmaceutical studies are those of Larsson and Lutz (2000) (neuropeptides including substance P) Hettiarachchi et al. (2001) (synthetic opioid peptides) Varesio et al. (2002) (amyloid-beta peptide) Zamfir and Peter-Katalinic (2004) (gangliosides) Peterson et al. (2002) (catecholamines and metanephrines) Cherkaoui and Veuthey (2002) (fluoxetine) and Smyth and Brooks (2004) (various lower molecular weight molecules including benzodiazepines, steroids, and cannabinols). 

Chan EC, Ho PC. 2000. High-performance liquid chromatog-raphy/atmospheric pressure chemical ionization mass spec-trometric method for the analysis of catecholamines and metanephrines in human urine. Rapid Commun Mass Spectrom 14 1959. 

Peterson ZD, Collins DC, BowerbankCR, Lee ML, Graves SW. 2002. Determination of catecholamines and metanephrines in urine by capillary electrophoresis-electrospray ioniza-tion-time-of-flight mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 776 221. 

Pheochromocytoma is a tumor of the adrenal medulla or sympathetic ganglion cells. The tumor secretes catecholamines, especially norepinephrine and epinephrine. The patient in the case study at the beginning of the chapter had a left adrenal pheochromocytoma that was identified by imaging. In addition, she had elevated plasma and urinary norepinephrine, epinephrine, and their metabolites, normetanephrine and metanephrine. 

Oxidation of fi-(4-Hydroxy-3-methoxyphenyl)ethylamines The j3-(4-hydroxy-3-methoxyphenyl)ethylamines [i.e. the catecholamine 03-methyl ethers, metanephrine (18) and normetanephrine (19)] are known to be among the major in vivo metabolites of... 

A sensitive spectrophotometric method based on the strong absorption of the aminochrome-sodium bisulfite addition products (see Section IV, F) at ca. 350 m/x. has been described recently by van Espen128and Oesterling and Tse 277-278 for determining total catecholamines. While not as sensitive as the fluorimetric procedures, this method is considerably more sensitive than the older colorimetric methods based on the visible absorption peak of the aminochromes. Also, it does not have many of the disadvantages (e.g. costly equipment and unstable blanks) often associated with fluorimetric techniques. The basic procedure can be satisfactorily applied to the differential determination of mixtures of adrenaline, noradrenaline, dopamine, metanephrine, and normetanephrine.178... 

Degradation of catecholamines The catecholamines are inacti vated by oxidative deamination catalyzed by monoamine oxidase (MAO), and by O-methylation carried out by catechol-O-methyl-transferase (COMT, Figure 21.15). The two reactions can occur in either order. The aldehyde products of the MAO reaction are axi dized to the corresponding acids. The metabolic products of these reactions are excreted in the urine as vanillylmandelic acid, metanephrine, and normetanephrine. 

Metanephrines represent metabolites of the catecholamines urinary levels are greater than total catecholamines but less than those of VMA. In tumors, variations in the metabolic pathways can cause an increase in the metanephrines alone. 

Since dopamine is present in sympathetic nervous tissue as a precursor of norepinephrine, and it has a separate metabolic pathway that yields homovanillic acid (HVA), tumors such as neuroblastomas may cause elevations of the urinary dopamine and its metabolite HVA. In some cases these elevations have been observed with normal VMA, total catecholamine, and metanephrine. Urinary HVA is usually normal in patients with phenochromocytoma. Increased HVA is found in special fluids of patients with Parkinson s disease treated with L-dopa. 

The most well known of the naturally occurring phenethylamine derivatives (Table I) are the transmitters of the sympathetic nervous system, epinephrine, norepinephrine, and dopamine. All these compounds are 3,4-dioxygenated in the aromatic nucleus and are collectively known as the catecholamines. Norepinephrine is the transmitter of most sympathetic postganglionic fibers, dopamine is the predominant transmitter of the mammalian extrapyramidal system and of several mesocortical and mesolimbic neuronal pathways, and epinephrine is the major hormone of the adrenal medulla (363). The literature that has accumulated on the action of these compounds in higher animals is enormous. Metanephrine and normetanephrine are known from animals as deactivated metabolites of epinephrine and norepinephrine that result from the action of the enzyme catechol O-methyltransferase (364). 

Biotransformations Epinephrine, like the other catecholamines, is metabolized by two enzymatic pathways COMT, which has S-adenosylmethionine as a cofactor, and MAO (see Figure 6.3). The final metabolites found in the urine are metanephrine and vanillylmandelic acid. [Note Urine also contains normeta-nephrine, a product of norepinephrine metabolism.]... 

Catecholamines can be variously oxidized or methylated. Extracellular epinephrine is O-methylated [via liver catechol-O-methyltransferase (COMT)] to 3-methoxyepinephrine (metanephrine) which can thence be oxidized [via monoamine oxidase (MAO)] to 3-methoxy-4-hydroxy-mandelic aldehyde and thence to 3-methoxy-4-hydroxyphenylglycol (MHPG) and 3-methoxy-4-hydroxy-mandelic. acid (VMA). Similarly, extracellular norepinephrine is O-methylated [via liver COMT] to 3-methoxynorepinephrine (normetanephrine) which can be oxidized [via MAO] to 3-methoxy-4-hydroxy-mandelic... 

Four catecholamines have been found in increased amounts in the urine after injury (W2), metanephrine, normetanephrine, iV-methyl-metanephrine, and 3-methoxytyramine. The JV-methylmetanephrine probably indicates an increased production of epinephrine (Cll). The increase in norepinephrine production is relatively greater than that of epinephrine, suggesting an active release of hormone from sympathetic nerve tissue as well as from the adrenal medulla. 

Catechol-0-methyltransferase (COMT) is responsible for the second major pathway of catecholamine metabolism, catalyzing 0-methylation of dopamine to methoxytyramine, norepinephrine to normetanephrine, and epinephrine to metanephrine. COMT is not present in monoamine-producing neurons, which contain exclusively MAO, but is present along with MAO in most extraneuronal tissue. The membrane-bound isoform of COMT, which has high affinity for catecholamines, is especially abundant in adrenal chromaffin cells. As a result of the preceding and other differences in the expression of metabolizing enzymes, catecholamines produced at neuronal and adrenal medullary locations follow different neuronal and extraneuronal pathways of metabolism (Figure 29-5). 

Findings that none of the traditionally used biochemical tests could reliably detect aU cases of pheochromocytomas led to recommendations that biochemical testing should include a combination of measurements of catecholamines and catecholamine metabolites. VMA is excreted in urine in large amounts, which makes measurement of this metabolite a simple, easy to implement, and time-honored test for diagnosis of pheochromocytomas. Numerous studies have now made it clear, however, that measurements of urinary VMA provide a relatively insensitive diagnostic test with limited value for initial testing for pheochromocytomas. Therefore the usual recommendation has been that biochemical testing should include measurements of urinary or plasma catecholamines and urinary metanephrines. 

The basis for the high diagnostic efficacy of plasma free metanephrines is explained by several factors (1) plasma free metanephrines are produced by metabolism of catecholamines within pheochromocytomas, a process that occurs continuously and independently of variations in catecholamine release by tumors (2) normally only small amounts of metanephrines are produced in the body, and these are relatively unresponsive to sympathoadrenal activation compared with the parent amines and (3) VMA and the metanephrines commonly measured in urine are different metabofites from the free metanephrines measured in plasma, and are produced in different parts of the body by metabolic processes not directly related to the tumor itself." ... 

Patterns of increases in plasma free metanephrines and catecholamines can also be useful for confirming pheo-chromocytomas in patients in whom initial tests of free metanephrines are positive but msufficiently elevated for a firm diagnosis. More specifically, patients with a pheochromocytoma usually have larger relative increases in metanephrines than of the parent catecholammes, whereas patients with false-positive results caused by sympathoadrenal activation usually have larger increases in catecholamines than metanephrines. 

Additional markers of catecholamine overproduction have been employed to improve the biochemical detection of neuroblastomas. Free dopamine may be abnormal in urine from neuroblastoma patients with VMA and HVA excretion. Combined testing for VMA, HVA, and dopamine may therefore improve tumor detection, and in 1993 an international consensus report on neuroblastoma diagnosis added dopamine to the Hst of acceptable measurements to document the adrenergic nature of the tumor. Plasma measurements of dopamine and L-dopa, the amino acid precursor of dopamine, may also have clinical value and allow the alternate use of plasma. Measurement of methylated metabolites, especially normetanephrine, has also been explored. When urinary normetanephrine, metanephrine, methoxytyra-mine, dopamine, norepinephrine, VMA, and HVA were measured, clinical sensitivity for detection of neuroblastomas was 97% to 100% when results of normetanephrine testing were coupled either with VMA in the infants or with HVA in children greater than age 1. Even with an extended panel of catecholamines and metabolite measurements, a low incidence of nonsecreting tumors continues to be identified and should be considered in the interpretation of a negative test result. 

Isolated deficiencies of MAO A and B are extremely rare and are associated with distinct clinical and neurochemical phenotypes. Deficiency of MAO A is associated with a behavioral disorder characterized by increased aggressiveness. Plasma and urinary levels of deaminated metabolites of catecholamines are severely decreased, whereas levels of normetanephrine and metanephrine are increased. An increased ratio of plasma normetanephrine to DHPG has therefore been proposed to provide a sensitive marker for the deficiency state. In contrast, deficiency of MAO B is associated with a mild phenotype, the only biochemical alteration is increased urinary excretion of phenylethyiamine. 

In contrast to the catecholamines, measurements of urinary metanephrines and VMA are still based in some routine laboratories on the early spectrophotometric assays developed by Pisano, Crout, and others in the late 1950s and early 1960s. Despite subsequent development of a variety of preanalytical cleanup and extraction procedures, these assays remain susceptible to analytical interference. They are also restricted to measurements in urine. Another limitation for spectrophotometric or fiuorometric assays of urinary metanephrines is that these methods do not allow separate (fractionated) measurements of normetanephrine and metanephrine. 

Interpretation of a biochemical test result as normal or abnormal depends on availability of valid reference intervals (see Chapter 16). For tests of a single analyte, such as VMA, it can be expected that at least 2.5% of patients without pheochromocytomas will have values for the analyte above the upper reference limit and 2.5% below the lower reference limit. Up to a 5% incidence of false-positive results might be expected for tests of pairs of analytes, such as norepinephrine and epinephrine in tests of urinary or plasma catecholamines or normetanephrine and metanephrine in tests of plasma free or urinary fractionated metanephrines. False-positive rates usually, however, tend to be higher than expected this is likely due to reduced control over sampling conditions and sources of interference or differences in clinical characteristics of reference and patient populations. 

Use of appropriately matched reference populations can be important for effective diagnosis of monoamine-producing tumors among different populations of patients tested for such tumors. Urinary and plasma levels of catecholamines and metanephrines show different ranges in hypertensives or hospitalized patients compared with norraotensive healthy volunteers, children compared... 

Reference intervals for plasma and urinary catecholamines and catecholamine metabolites also differ according to sex and age. Females have lower plasma concentrations of epinephrine and metanephrine than males. Similarly, 24-hour urinary outputs of catecholamines and metanephrines are lower in women than men for epinephrine this difference remains significant when values are normalized for creatinine excretion Plasma levels of norepinephrine and normetanephrine increase with advancing age in adults, whereas plasma levels of epinephrine and metanephrine are little affected. Age-related increases in 24-hour urinary outputs of norepinephrine and normetanephrine have also been reported,but not consistently by all studies. In general, the influences of age... 

Normetanephrine andmetanephrine are metabolic products of norepinephrine and epinephrine, respectively, and are formed by the action of catechol-0-methyltransferase without deamination. As a result of active neuronal reuptake and deamination of norepinephrine, normetanephrine normally represents <5% of the total norepinephrine excretion products in urine. Metanephrine, however, even with its lower urinary concentration relative to normetanephrine, represents a major excretion product of epinephrine. The metanephrines are excreted in both conjugated and unconjugated forms. Unlike the catecholamines, total metanephrine excretion is not significantly influenced by diet. As a result, the total metanephrines are routinely measured after acid hydrolysis or sulfatase pretreatment. 

Eisenhofer G, Rundquist B, Aneman A, Friberg P, Dakak N, Ropin IJ, et al. Regional release and removal of catecholamines and extraneuronal metabolism to metanephrines. J Clin Endocrinol Metab 1995 80 3009-17. 

Feldman JM. Falsely elevated urinary excretion of catecholamines and metanephrines in patients receiving labetalol therapy. J Clin Pharmacol 1987 27 288-92. 

Munion CL, Seaton JF, Harrison TS. HPLC for urinary catecholamines and metanephrines with alpha-methyldopa. J Surg Res 1983 35 507-14. 

Parker NC, Levtzow CB, Wright PW, Woodard LL, Chapman JF. Uniform chromatographic conditions for quantifying urinary catecholamines, metanephrines, vanillylmandelic add, 5-hydroxyindoleacetic acid, by liquid chromatography, with electrochemical detection. Clin Chem 1986 32 1473-6. 

Sawka AM, Jaeschke R, Singh RJ, Young WF, Jr. A comparison of biochemical tests for pheochromocytoma measurement of fractionated plasma metanephrines compared with the combination of 24-hour urinary metanephrines and catecholamines,... 

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