Metanephrines

As with the catecholamines, fluorescence methods have also been reported for urinary metanephrine analysis. Fluorescent derivatization of the metanephrines NM and MN by chemical oxidation was based on modification of the trihydroxyindole reaction used for catecholamines. The individual metanephrines were measured following chromatographic separation and fluorescent derivatization or through the formation of differential fluorescent compounds by oxidation at different pH levelsSince the stability of the fluorescent products was variable, with some products decomposing within 10 min,this method has limited application in current practice. Other early methods for analysis of NM and MN included electrophoresis and paper and thin-layer chromatography. These assays were technically complex and had poor analytical sensitivity. 

Measurement of total urinary metanephrines using spectrophotometric methods has also been reported. After sample hydrolysis and isolation of metanephrines by ion-exchange chromatography, the hydroxyl- and amino-containing side-chains were oxidatively cleaved by periodate. Oxidation of both NM and MN resulted in the formation of a common end product, vanillin, which was measured spectrophotometricafly at 360 nm. This method was, however, susceptible to interferences from other compounds such as methylglucamine and the 4-hydroxylated metabolite of propranolol. Furthermore, the method did not differentiate between NM and MN, and results were only semi-quantitative for grossly increased concentrations of total metanephrines.  

Radioenzymatic methods had also been used to quantify urinary NM. Phenyletha-noleamine-A -rnethyltransferase and H iS -adenosylmethionine convert NM to its [ H]N-methylated derivative, The main advantages of this method were its high 

---

More specific laboratory tests are used to diagnose secondary hypertension. These include plasma norepinephrine and urinary metanephrine levels for pheochromocytoma, plasma and urinary aldosterone levels for primary aldosteronism, and plasma renin activity, captopril stimulation test, renal vein renins, and renal artery angiography for renovascular disease. 

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. 

Acebutorol, arotinolol, atropine, bupivacaine, clorprenaline, denopamine, eperisone, epinastine, etirefline, fenoterol, homatropine, ketamine, metanephrine, metoprolol, mexiletine, nicardipine, oxyphencyclimine, phenylephrine, pindolol, primaquine, promethazine, sulpiride, ter-butaline, tolperizone, trihexyphenidyl, trimebutine, trimetoquinol, trimipramine, verapamil 81... 

Newer MAOI drugs are selective for the MAO-A subtype of the enzyme, and are less likely to interact with foods or other drugs. Monoamine oxidase (MAO) inactivates monoamine substances, many of which are, or are related to, neurotransmitters. The central nervous system mainly contains MAO-A, whose substrates are adrenaline (epinephrine), noradrenaline (norepinephrine), metanephrine, and 5-hydroxyti7ptamine (5-HT), whereas extra-neuronal tissues, such as the liver, lung, and kidney, contain mainly MAO-B which metabolises p-phenylethylamine, phenylethanolamine, o-tyramine, and benzylamine. 

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... 

The mechanisms of these interesting oxidations require further study, and attempts should be made to isolate the intermediate aminochromes (or a simple derivative, such as the semicarbazone). However, it is difficult to escape the conclusion that aminochromes can be readily obtained by the oxidation of compounds such as metanephrine and normetanephrine. 

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. 

Gamache, P. H., Kingery, M. L., and Acworth, I. N. (1993). Urinary metanephrine and norme-tanephrine determined without extraction by using hquid chromatography and coulometric array detection. Clin. Chem. 39 1825-1830. 

The following were isolated largely by chromatography and identified by spectral properties and by comparison with known bases N-formylnormacromerine, metanephrine, V-methylmetanephrine, syne-phrine, N- methyltyramine, V-methyl-4-methoxy- -phenethylamine, iV-methyl-3,4-dimethoxy-/3-phenethylamine, tyramine, hordenine, macromerine, and normacromerine (37). 

On-chip chiral separation of enantiomers of adrenaline, noradrenaline, and dopamine was achieved using a mixture of carboxymethyl-p-cyclodextrin (CMCD) and a polyamidoamine dendrimer (Starburst) [120], On-chip chiral separation of enantiomers of homovanillic acid, DOPA, cDOPA, methoxy-tyramine (MT), metanephrine, and normetanephrine was achieved using a mixture of 18-crown-6-ether and carboxymethyl-P-cyclodextrin [120],... 

Coryphantha missouriensis (Sweet) metanephrine, N-methylmetaneph-rine, nor-Macr, Macr, N-formyl-nor-Macr Tyr, N-Me-Tyr, Hord, N-Me-Homova 72... 

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). 

---