Synthesis of a biochemically important aldehyde, 3,4- dihydroxyphenylacetaldehyde

Article abstract of DOI:10.1006/bioo.1998.1087

3,4-Dihydroxyphenylacetaldehyde (DOPAL) is an important precursor of the major brain metabolites of dopamine, 3,4-dihydroxyphenylacetic acid and 4- hydroxy-3-methoxyphenylacetic acid. A new method for the synthesis of DOPAL from piperonal is described. We report for the first time the physical and chemical characteristics of DOPAL. Its importance for research in Parkinson's disease and Alzheimer's disease is discussed.

Full text of DOI:10.1006/bioo.1998.1087

BIOORGANIC CHEMISTRY 26, 45–50 (1998)
ARTICLE NO. BH981087
Synthesis of a Biochemically Important Aldehyde,
3,4-Dihydroxyphenylacetaldehyde
Shu Wen Li,*,†,1 Vincent T. Spaziano,* and William J. Burke†
*Department of Chemistry and †Department of Neurology, Veterans Administration Medical Center,
and Saint Louis University Health Sciences Center, St. Louis, Missouri 63110
Received December 23, 1997
3,4-Dihydroxyphenylacetaldehyde (DOPAL) is an important precursor of the major brain
metabolites of dopamine, 3,4-dihydroxyphenylacetic acid and 4-hydroxy-3-methoxy-
phenylacetic acid. A new method for the synthesis of DOPAL from piperonal is described.
We report for the first time the physical and chemical characteristics of DOPAL. Its importance
for research in Parkinson’s disease and Alzheimer’s disease is discussed.
1998 Academic Press
INTRODUCTION
3,4-Dihydroxyphenylacetaldehyde (DOPAL) is formed by oxidative deamination
of dopamine (DA) by monoamine oxidase (MAO) (1, 2). In tissues this aldehyde
is either enzymatically oxidized to 3,4-dihydroxyphenylacetic acid or reduced to
3,4-dihydroxyphenylethanol (DOPET) (2, 3). The synthesis of DOPAL is important
for several reasons (see Fig. 1). First, a standard will allow an accurate measurement
in human and animal tissues. Second, the human brain, in a number of disorders
such as Parkinson’s disease and Alzheimer’s disease, is studied primarily using
postmortem tissue. Deficits in catecholamines, including DA, are thought to play
a role in these disorders (4, 5). DOPAL is the precursor of other major brain
metabolites of DA, including 3,4-dihydroxyphenylacetic acid and 4-hydroxy-3-meth-
oxyphenylacetic acid or homovanillic acid (6). Carlsson and Winblad (7) have
suggested that in postmortem brain the sum of a catecholamine plus its major
metabolites is a better index of the level of the catecholamine in brain at the time
of death than the catecholamine alone. This is supported by the findings of Warsh
et al. (8) that catecholamines are metabolized in the brain after death. The measure-
ment of DOPAL will therefore contribute to a more accurate estimation of premor-
tem levels of DA in postmortem human brain. In addition, because the enzymes
involved in metabolism of DOPAL (aldehyde reductase, aldehyde dehydrogenase,
and catechol-O-methyltransferase) either are in very low amounts (9) or are extra-
neuronal (10–12), DOPAL may be an index of the intraneuronal metabolism of
newly formed DA (13).
Finally, DOPAL is the initial MAO-B metabolite of dopamine. Blashko (1) has
suggested that aldehyde metabolites generated by the action of MAO on amines
1 To whom correspondence and reprint requests should be addressed.
45
0045-2068/98 $25.00
Copyright 1998 by Academic Press
All rights of reproduction in any form reserved.

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LI, SPAZIANO, AND BURKE
FIGURE 1
may be toxic to cells in which they are formed. Markey et al. (14) have shown that
the exogenous toxin 1-methyl-4-phenyl-1,2,4,6-tetrahydropyridine, which causes a
form of Parkinson’s disease, becomes toxic to neurons only after its oxidation to
the pyridinium form by MAO-B. We have previously synthesized 3,4-dihydroxyphe-
nylglycolaldehyde (DOPEGAL), the MAO-A metabolite of noradrenaline (NA)
and adrenaline (15). DOPEGAL is a major NA metabolite in animal and postmor-
tem human tissue (16, 17). DOPEGAL, but not other noradrenaline metabolites,
is toxic to cultured PC-12 cells (18) and triggers apoptotic cell death (19), the form
of neuronal death found in Alzheimer’s disease (20) and Parkinson’s disease (21).
Attempts have been made to synthesize pure DOPAL (22–25). However, none
of the methods provide detailed procedures and physical or chemical characteriza-
tion of DOPAL other than its molecular weight by GC–MS. Such a method is
important because, in order to study a potential role for DOPAL in neuron death
in Alzheimer’s disease and Parkinson’s disease, it is necessary to develop a reliable
method for synthesis and purification of large amounts of chemically pure DOPAL.
The chemically pure compound is required for in vitro and in vivo tests of DOPAL
toxicity as well as for quantification of DOPAL in brain neurons affected in Parkin-
son’s disease and Alzheimer’s disease (21, 26–28).
In this paper we describe a new synthetic method for DOPAL and its physical
characterization. The successful synthetic route uses commercially available pipero-
nal as outlined in Scheme 1. Conversion of 2 to the ethyl glycidate (3) was accom-
plished by reaction with ethyl chloroacetate in the presence of sodium ethoxide.

SYNTHESIS OF 3,4-DIHYDROXYPHENYLACETALDEHYDE
47
SCHEME 1. (a) Ethyl chloroacetate, NaOC2H5; (b) NaOH, ethanol; (c) acetic acid, benzene; and
(d) BCl3/S(CH3)2 , 1,2-dichloroethane.
Hydrolysis of 3 with sodium hydroxide gave the sodium salt (4) which upon treat-
ment with acetic acid in benzene generated the aldehyde group to afford 5. Final
deprotection of the methylenedioxy group with boron trichloride–methyl sulfide
complex gave crude 1, which was purified by chromatography over silica gel.
EXPERIMENTAL
Melting points were determined on a Fisher–Johns melting point apparatus and
uncorrected. NMR spectra were recorded in CDCl3 or DMSO-d6 on a Varian
Gemini 300 spectrometer with Me4Si as an internal standard. Field strength of
various proton resonances is expressed as s, d, t, q, the center of which is given.
IR spectra were recorded on a Perkin–Elmer Paragon 1000 FT-IR spectrometer.

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LI, SPAZIANO, AND BURKE
Ultraviolet spectra were recorded on a Hewlett–Packard 8452A diode array spectro-
photometer. GC–mass spectrometry was obtained using an HP 5890 Series II gas
chromatograph coupled with a HP 5971 mass selective detector. The gas chromato-
graph was equipped with Alltech Econo Cap SE-45 capillary column (30 m ϫ 0.25
mm i.d. ϫ 0.25 Ȑm film thickness). Molecular masses were determined via a VG70-
250 SEQ hybrid-tandem spectrometer. Column chromatography was performed on
silica gel 60 (70–230 mesh ASTM) and TLC analysis on silica gel 60 F254 precoated
plates (layer thickness 0.2 mm).
Ethyl-3-(3,4-Methylenedioxyphenyl) Glycidate (3)
To 65 ml of absolute alcohol was added 3.08 g (0.134 gram atom) of sodium at
room temperature. The reaction mixture was heated under reflux until the solid
sodium disappeared and then cooled to 0–5ЊC. A mixture of piperonal (2) (17.8 g,
0.119 M) and redistilled ethyl chloroacetate (15.93 g, 0.13 M) was added to the
above sodium ethoxide solution at Ϫ5ЊC during 1 h. The reaction mixture was
stirred at 0ЊC for 1 h then allowed to warm to room temperature overnight and
poured into a mixture of ice (100 g) and acetic acid (2 ml). The mixture was
extracted with dichloromethane (50 ml ϫ 4) and the extracts were washed with
water and dried over anhydrous sodium sulfate. Removal of the solvent gave a
crude product which was purified by chromatography on a silica gel column eluting
with 50% dichloromethane in hexane. The desired fractions were identified with
TLC, pooled, and evaporated to afford an oil, 18.8 g (67%) of 3; NMR (CDCl3), ͳ
6.84 (dd, 1H, aromatic), 6.78 (d, 1H aromatic), 6.70 (d, 1H aromatic), 5.97 (s, 2H,
CH2), 4.28 (q, 2H, CH2), 4.02 (d, 1H, J ϭ 2 Hz, CH), 3.46 (d, 1H, J ϭ 2 Hz, CH),
1
1.30 (t, 3H, CH3); IR (nujol) 1740 (CO), 931 (OCH2O), 1235 (–O–), cmϪ ; MS
(GC–MS), calcd for C12H12O5: 236.00. Found: 236 (Mϩ).
Sodium 3-(3,4-methylenedioxyphenyl) Glycidate (4)
To a solution of 3 (11.48 g, 0.049 M) in 35 ml of ethanol was added 25 ml of 4
N NaOH and 10 ml of water. The reaction mixture was stirred at room temperature
overnight. The solid which separated was collected, washed with ethanol, and dried
over P2O5 in a desiccator: yield 9.1 g (81%) of 4, mp Ͼ300ЊC. The dried salt was
used without further purification.
3,4-Methylenedioxyphenylacetaldehyde (5)
To a suspension of 4 (8.87 g, 0.038 M) in 100 ml of benzene was added 10 ml of
acetic acid. The suspension was heated under reflux for 4 h. The benzene solution
was washed with saturated sodium bicarbonate solution then water and dried over
anhydrous sodium sulfate. Removal of the benzene gave a crude compound which
was purified by chromatography on a silica gel column with 30% of dichloromethane
in hexane as the eluting solvent. The desired fractions were identified with TLC,
pooled, and concentrated: yield 3.6 g (57%) of 5, mp 60–62ЊC; NMR (CDCl3), ͳ
9.70 (t, 1H, CHO), 7.22 (dd, 1H, aromatic), 7.19 (d, 1H aromatic), 6.85 (d, 1H,
aromatic), 6.10 (s, 2H, CH2), 3.59 (d, 2H, CH2); IR (nujol), 1720 (CO), 927
1
(–OCH2O–) cmϪ ; MS (GC–MS), calcd for (C9H8O3): 164.00. Found: 164 (Mϩ).

SYNTHESIS OF 3,4-DIHYDROXYPHENYLACETALDEHYDE
49
3,4-Dihydroxyphenylacetaldehyde (1)
To a solution of 5 (560 mg, 3.42 mM) in 15 ml of 1,2-dichloroethane was added
3.5 ml of 2 M boron trichloride–methyl sulfide complex solution in dichloromethane.
The reaction mixture was heated under reflux for 24 h and then cooled to room
temperature. Ten milliliters of water was added and the mixture was stirred for
1 h at room temperature. The organic layer was separated and the water layer was
extracted with 1,2-dichloroethane (15 ml ϫ 3). The combined extracts were washed
with water and dried over anhydrous sodium sulfate. Removal of the solvent gave
the crude product which was purified by chromatography on a silica gel using 2%
methanol in dichloromethane as the eluting solvent. The desired fractions were
identified with TLC, pooled, and concentrated to afford 75 mg (14%) of 1: NMR
(DMSO-d6) ͳ 9.67 (t, 1H, CHO), 7.27 (dd, 1H, aromatic), 7.20 (d, 1H, aromatic),
6.90 (d, 1H, aromatic), 3.50 (d, 2H, CH2); mp 110–112ЊC; IR (nujol), 3425 (OH),
1
1690 (CHO) cmϪ ; UV (methanol) ␭ max 228 nm (␧ ϭ 9173), 262 nm (␧ ϭ 8181);
MS (CI), calcd for C8H8O3: 152.00. Found: 153 (MHϩ). Anal. Calcd for C8H8O3H2O:
C, 56.45%; H, 5.92%. Found: C 56.18%; H, 6.18%.
ACKNOWLEDGMENTS
This work was supported by a grant from the Veterans Affairs Merit Review program. We express
our thanks to Dr. F. A. Bancsach and Dr. Betsy Ann, University of South Alabama, for aid in molecu-
lar masses.
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