SIROSENOH,C. R. CREVELING, SIDNEY UDENFRIENDA N D BERNHARD WITKOP
6236
Vol. SI
[CUNTRIBCTION FROM THE NATIONAL INSTITUTE O F h T H R I T I S AND METABOLIC DISEASES, NATIONAL INSTITUTES OP IIEALTH, PUBLIC HEALTH SERVICE]
Chemical, Enzymatic and Metabolic Studies on the Mechanism of Oxidation of Dopamine BY SIROSENOII,~ CYRUSR. CREVELING, SIDNEYUDENFRIEND AND BERNHARD WITKOP RECEIVED MAY15, 1950 N-Carbobeiiz~-loxy-3,4-dihydrox)-pl1eiietliylan1ine-~,~-H~ suffers no loss of tritium activity when subjected to oxidatioti to the quinone, to 1,4-additionreactions involving catalysis by boron trifluoride etherate or to the conditions of decarbobenzyloxylation in 48% hydrobromic acid. N-Carbobenzyloxy-6-methoxydopaminequinone-p,p-H3 (XI) shows only slight tritium lability a t room temperature which increases at higher pH. The exchange of tritium in a comparable intermediary quinone of dopamine (111, IV) would be too slow to be detectable in the enzymatic synthesis of norepinephrine which ha5 been studied with a number of tissues using dopamine labeled with C14 and H3. The responsible enzyme ("doparnine 0oxidase") effects conversion to norepinephrine with a H3/CI4 ratio approximating half of that of the starting material. The retention of the original H3/C1*ratio in other experiments led to the discovery of the isomeric 2,4,5-trihydroxyphenethylamine (11),isogruphic with norepinephrine in more than ten different solvent systems. It is formed from dopamine by oxidation, autoxidation or metabolic processes. Whether nuclear hydroxylation proceeds enzymatically and whether it and thc IV must be left open, until direct studies with 0 1 8 biogenesis of norepinephrine pass through quinoid intermediates I11 -4 become practicable.
The biosynthesis of the endogenous sympathetic hormone norepinephrine (V), which starts from tyrosine, is generally believed to involve hydroxylation of the side chain of dopamine (I)4 in the sequence of reactions (Chart I) .5,6 This conversion
:x3 c
H
c H,S H . . ,
,,
,
,,
c
,,
, H , O p c r ICI12pI'IIz
-
6 4
former (11) could also arise by pathway A, aiialogous to D, i e., direct hydroxylation, of I . The fiquinone methine IV, a t least enzymatically, need not be formedvia the o-quinone I11 (pathway B), but could arise directly by 1,6-abstraction of hydrogen (pathway C). An initial 9-quinol undergoing an allylic shift, such as observed in the easy rearrangement of VI + VI1 (X = Br),8is possible in a highly hindered p substituted phenol but unlikely in a catechol. 0 1I
0
-L*.4
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..
xx i'
1 ,
L.+,
I:
!
y
~ k ? ~ i n . , ! ''dujiItIi!iIIL ~ ,
s-uxidase"
110~~~L~u~*C~~,,,,, J A
110'
11 Cliart I.-Enzymatic
,' x
It-(
,
1
--
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and non-enzymatic conversions of dopamine.
has been demonstrated in isolated tissues marly times. However, i t has not been possible so far to obtain enzyme preparations potent enough to permit studies on the mechanism of the reaction, e.g., with 0l8. The latter could enter directly into the benzyl position of the side chain (pathway D)'I or, conceivably, could first dehydrogenate I to the quinone I11 (pathway B) which itself, or as the tautomeric quinone methine IV, is capable of l,&- or 1,6addition of water1 to yield either 2,4,5trihydroxyphenethylamine (11) or norepinephrine (V). The (1) Oxidation Mechanisms. XXIV. Preceding paper cf. THIS 81, 6231 (1959). (2) Visiting Scientist of t h e USPIIS on leave of absence from t h e Institute of Food Chemistry and Osaka City University, Japan. (3) S. Udenfriend and J. B. Wyngaarden, Biochirn. Bioghys. A
