Oxidation and degradation products of papaverine. Part I: Gadamer and Schulemann's papaverinol synthesis revisited

Article abstract of DOI:10.1002/1521-4184(200204)335:4<167::AID-ARDP167>3.0.CO;2-O

In 1915 Gadamer published in this journal [1] a procedure for the synthesis of papaverinol 2 from papaverine 1 in excellent yield. However, he did not investigate the formation of a violet fluorescence produced upon crystallization of papaverinol 2 from ethanol. The compound responsible for this fluorescence was isolated and identified as the yet unknown quaternary ammonium ion 4, a 6a, 12a-diazadibenzo[a,g]fluorenylium derivative. The isolation of 4 and its structure determination by spectroscopic methods are described. However, its formation mechanism is unknown.

Full text of DOI:10.1002/1521-4184(200204)335:4<167::AID-ARDP167>3.0.CO;2-O

Arch. Pharm. Pharm. Med. Chem. 2002, 4, 167–169
Oxidation and Degradation Products of Papaverine 167
Tadeusz W. Hermanna,
Oxidation and Degradation Products of Papaverine.
Part I: Gadamer and Schulemann’s Papaverinol
Synthesis Revisited
Ulrich Girreserb,
Pawel Michalskia,
Karolina Piotrowskaa
aDepartment of Physical
Chemistry,
K. Marcinkowski University of
Medical Sciences, Poznan´ ,
Poland
bDepartment of
Pharmaceutical Chemistry,
Christian-Albrechts-University,
Kiel, Germany
In 1915 Gadamer published in this journal [1] a procedure for the synthesis of pa-
paverinol 2 from papaverine 1 in excellent yield. However, he did not investigate the
formation of a violet fluorescence produced upon crystallization of papaverinol 2
from ethanol. The compound responsible for this fluorescence was isolated and
identified as the yet unknown quaternary ammonium ion 4, a 6a,12a-diazadibenzo-
[a,g]fluorenylium derivative. The isolation of 4 and its structure determination by
spectroscopic methods are described. However, its formation mechanism is un-
known.
Keywords: Papaverine oxidation; Diazafluorene derivative; Hg2+ cations; Column
chromatography; Structure evaluation
Received: September 19, 2001 [FP632]
Introduction
Synthesis
Mercury(II) acetate was used to oxidize the base 1. Mer-
Papaverinol 2 and papaveraldine 3 are papaverine oxi-
dation products, which contaminate authentic samples
and are also found in its injection solutions [2–4].
Papaverine N-oxide and 6,7-dimethoxyisoquinoline N-
oxide were identified as products of papaverine photo-
oxidation [5]. Polarography provided evidence that
papaveraldine 3 is further oxidized to a brown compound
of polymeric nature under the influence of sun- and day-
light [6, 7]. Different methods were used for synthesis of
papaverinol from papaverine [1, 2, 8]. Gadamer and
Schulemann [1] oxidized papaverine 1 to papaverinol 2
in excellent yield (93.6 %) [2].They used mercury(II) ace-
tate [1] as oxidant. However, they did not investigate the
side products (6.4 %), the structures of which have not
been analyzed up to now. We focussed our attention on
the major side product detected by virtue of its violet-
blue fluorescence which developed on crystallization of
papaverinol 2 from ethanol, and is also observed on
washing of HgS precipitated upon addition of thioacet-
amide to the reaction mixture in order to separate all the
mercury salts present.Therefore, the goal of our experi-
ments was the isolation and identification of the com-
pound responsible for the violet-blue fluorescence men-
tioned above.
cury(I) acetate was filtered off and the excess of Hg2+
ions was removed.Then the filtrate was alkalized and ex-
tracted with chloroform.The extract was first desiccated
and then evaporated to dryness. Papaverinol 2 was re-
crystallized from ethanol. The filtrate was evaporated to
dryness and the residue was separated by column chro-
matography. After careful elution of any residual pa-
paverinol with a mobile phase consisting of chloroform-
methanol (19:1 and 18:2), a mixture of chloroform and
methanol (1:1, v/v) was used to isolate the band of violet
fluorescence of the compound under investigation,
which turned out to be 13-(3,4-dimethoxyphenyl)-
2,3,8,9-tetramethoxy-6a,12a-diazadibenzo[a,g]fluoren-
ylium chloride 4. The above compound base (mp
228–232°C) was identified by means of UV/vis, fluores-
cence, 1H and 13C NMR spectroscopy, and mass spec-
trometry. It was also crystallized as chloride (mp
226–238°C, decomp.). The above crystallization proce-
dure permits removal of a red-colored impurity. Other
salts of 4, such as the picrate (mp 299–302°C) and the
perchlorate (mp 324–327°C), can also be obtained.
Spectroscopic evaluation of the structure of 4
The UV/vis and fluorescence spectrum (in methanol)
confirmed the optical properties, 4 showing the absorp-
tion maximum of longest wavelength at 393 nm. The
fluorescence maximum was observed at 430 nm (uncor-
rected), when the excitation wavelength was set to either
Correspondence: T. W. Hermann, Department of Physical
Chemistry, K. Marcinkowski University of Medical Sciences, 6
Swiecickiego Street, 60 781 Pozna´n, Poland. Fax:
+48 61 865 70 05, e-mail: hermann@mail.usoms.poznan.pl.
© WILEY-VCH Verlag GmbH, 69451 Weinheim, 2002
0365-6233/04/0167 $ 17.50+.50/0

168 Hermann et al.
Arch. Pharm. Pharm. Med. Chem. 2002, 335, 167–169
nately, the 1H resonance of the aromatic protons of the
3,4-dimethoxyphenyl group are partially overlapping in
all of the solvents tested (perdeuterated acetone, chloro-
form, and dimethyl sulfoxide), making the assignment of
these signals arbitrary.
Discussion
On Gadamer and Schulemann's procedure [1] papaver-
ine 1 is mainly oxidized (93.6 %) to papaverinol (6,7-
dimethoxyisoquinoline-1-(3,4-dimethoxyphenyl)-1-car-
binol). Among the remaining 6.4 %, an unknown oxida-
tion product 4 has been identified as the major constitu-
ent. The mechanism of its formation is still unknown.
However, it can be obtained neither from papaverinol 2
nor from papaveraldine 3 with mercury(II) acetate. It is
formed as a minor product only (0.6 %) of a parallel reac-
tion of papaverine 1 oxidation with mercury(II) acetate
according to the major pathway of Gadamer and Schule-
mann's synthesis leading to papaverinol (94 %) which is
further oxidized to papaveraldine 3 via a very minor se-
quential reaction (< 1 %) (Scheme 1). We have also
proved that an equimolar mixture of papaverine 1 and
1-methyl-6,7-dimethoxyisoquinoline can be oxidized to 4
with mercury(II) acetate, but the yield is still very low
(0.6 %), identical to that yield of the reaction without
1-methyl-6,7-dimethoxyisoquinoline. It should also be
mentioned that the presence of 1-methyl-6,7-dimeth-
oxyisoquinoline in a mixture with either papaverinol 2 or
papaveraldine 3 does not generate at all the new com-
pound 4 with mercury(II) acetate. It seems to us that a
radical mechanism may be involved in the above reac-
tion of 4 formation. It is found in the literature [9] that pa-
paverine possesses catalytic properties in the indigo
carmine oxidation reaction with hydrogen peroxide. It is
postulated that free radicals can be formed from
papaverine and could be responsible for its catalytic
properties. Furthermore, hydrogen atoms of the
papaverine methylene group presumably form free radi-
cals, which activate oxygen in hydrogen peroxide [10].
Therefore, condensation of a free radical of papaverine
with 6,7-dimethoxyisoquinoline, formed from another
molecule of the papaverine free radical, and subsequent
dehydrogenation would afford the base of the isolated
product 4.The above radical pathway is only a hypothe-
sis, requiring further investigation and is therefore not
depicted in Scheme 1.The parent heterocycle was pre-
pared according to Brown et al. from 2-bromopyridine
and 2-bromomethyl pyridine [11, 12]. All the physical
chemical properties of 4, i.e. its solubility in organic sol-
vents as a base and its spectroscopic data, are in ac-
cordance with the postulated structure.
Scheme 1
285 or 395 nm.The appearance of these spectra, howev-
er, resembled those observed for papaverine hydrochlo-
ride (242, 56742; 280, 7539; 313, 5118, and 329, 4792,
λmax [nm] and ε [mol–1 cm–1 L], respectively) being
bathochromically shifted, suggesting that a dimethoxy
substituted isoquinoline moiety is still present. However,
a hyperchromic effect is observed for the absorption
maximum at the longest wavelength pointing towards
the presence of an additional isoquinoline moiety in
structure 4.
The electron impact mass spectrum did not allow the de-
termination of the molecular mass due to strong pyroly-
sis in the inlet system, thus the electrospray technique
(ESI, pos.ion) was used, affording a molecular ion at m/z
525, and an isotope pattern (m/z 525:526:527) =
100:36.5:8.5 %. Interestingly, this ion was obtained as
well when dissolving 4 in D2O. Enhancement of the ion
yield by addition of appropriate buffers was not ob-
served, evidencing that the compound under investiga-
tion already contains a quaternary nitrogen atom and is
not ionized by the solvent system. Furthermore MS/MS
experiments in the ion trap of the mass spectrometer in-
dicated a very stable and presumably aromatic structure
as only small neutral fragments (CH4, H2O) were lost,
with the major fragmentation series at m/z 525 → 509 →
493 → 475.
NMR spectroscopy confirmed the mass spectrometric
data, affording the molecular formula (C31H29O6N2)+Cl–,
(isotopic pattern calculated 100:36.1:7.5 %), which was
confirmed by elemental analysis (Found/Calcd, C 61.27/
66.37, H 5.425/5.210, N 4.655/4.990, Cl 6.23/6.32). All
the hydrogen and carbon atoms resonate at different fre-
quencies, no exchangeable protons were present in 4.
The structure of 4 was assembled using two-dimension-
al NMR techniques such as H,H- and C,H long range
correlation. The assignments of the H and 13C reso-
Further investigations will concentrate on a synthetic ap-
1
nance are given in the experimental section. Unfortu-
proach to 4 in order to explain the mechanism of its for-

Arch. Pharm. Pharm. Med. Chem. 2002, 335, 167–169 Oxidation and Degradation Products of Papaverine 169
ethanol filtrate was combined with the previously obtained
ethanol solutions and was evaporated to dryness in vacuo and
the residue was purified by the column chromatography proce-
dure mentioned above. Mobile phases consisting of chloro-
form, chloroform-methanol (19:1, 18:2, and 1:1, v/v) were
used, respectively. The chromatographic fractions with pure
compound 4 were combined and evaporated to dryness in
vacuo and the rose-white residue (25 mg, 0.64 %) was
analyzed by spectroscopic means specified above. The chlo-
ride of 4 could also be crystallized from methanol with a few
drops of 10 % HCl and a few drops of benzene or toluene to pro-
duce a white turbidity.The mixture was first cooled to room tem-
perature and then in a refrigerator to obtain thin, long white-
yellowish needles (12 mg) which decompose when melted at
226–236°C.Compound 4 develops an intensive green color on
reaction with concentrated sulfuric acid.
mation and to obtain sufficient quantities to evaluate the
pharmacological properties of this interesting structure.
Acknowledgement
Prof. T. W. Hermann appreciates the scholarship obtained for
one month from the Christian-Albrechts-University at Kiel and
all excellent research facilities provided at the Departments of
Pharmaceutical Chemistry (Prof. B. Clement) and Pharmaceu-
tical Technology and Biopharmacy (Prof. B. W. Müller).
Experimental section
1H NMR (300 MHz, DMSO-d6): δ ppm 3.44, 3.81, 3.93, 3.94,
4.04, 4.24 (6 × s, 6 × 3H, 8-, 3´-, 9-, 4´-, 3-, 2-OCH3), 7.08 (s,
1H, H-7), 7.40 (m, 2H, H-5´, H-6´), 7.45 (m, 1H, H-2´), 7.60 (d,
J = 6.0 Hz, 1H, H-11), 7.67 (s, 1H, H-10), 7.92 (s, 1H, H-4),
7.96 (d, J = 6.5 Hz, 1H, H-5), 8.20 (s, 1H, H-1), 8.25 (d, J =
6.5 Hz, 1H, H-6), 9.26 (d, J = 6.0 Hz, 1H, H-12).
Melting points were determined on a Boëtius microscope and
are not corrected. EI (70 eV) mass spectra were recorded on a
HP-MS engine 5989 A and are consistent with the assigned
structure 4. ESI mass spectra and MSn experiments were per-
formed with
a Bruker Esquire~LC mass spectrometer.
300 MHz 1H- and 75 MHz 13C NMR spectra were recorded on a
Bruker ARX 300 spectrometer. Chemical shifts are given in
ppm and TMS was used as an internal standard for the spectra
obtained in DMSO-d6 and CDCl3.The IR spectrum was record-
ed as KBr pellet on a Perkin Elmer 16 PC FT-IR instrument. A
fluorescence spectrometer Perkin Elmer LS 50 B was used.
UV/vis spectra were recorded on a Specord M-40 (Carl Zeiss,
Jena) and 4 showed maxima at 242, 280, 374, and 393.5 nm
(ε [mol–1 cm–1 L], 41480, 24383, 11883, 11883, respectively).
Column chromatography was performed on aluminium oxide
(ICN Alumina B-Super I), whose absorption activity III was ob-
tained by addition of 7 % of water and was employed for the iso-
lation of 4 from the reaction mixture.The purity of the fractions
was monitored by TLC on silica gel plates (Polygram® 40 ×
80 mm SIL G/UV254) with chloroform-methanol (18:2, v/v).
Compound 4 was visualized on the column and the TLC plates
with a Hanau Fluotest Heraeus lamp at emissions at 254 and/or
366 nm. It shows a blue fluorescence when excited at 366 nm.
13C NMR (75 MHz, DMSO-d6): δ ppm 54.82, 55.78, 55.83,
55.88, 56.10, 56.65 (8-, 3´-, 9-, 4´-, 3-, 2-OCH3), 102.97 (C-1),
104.74 (C-7), 109.14 (C-4), 109.21 (C-10), 111.97 (C-13b),
113.02 (C-2´), 114.59 (C-5´), 115.55 (C-6c), 116.84 (C-11),
117.14 (C-1´), 117.71 (C-13), 118.69 (C-5), 119.86 (C-6),
120.70 (C-12), 122.79 (C-10a), 124.99 (C-6´), 125.26 (C-6b),
127.72 (C-4a), 128.21 (C-13a), 149.74 (C-8), 150.00 (C-3´),
150.83 (C-9), 151.01 (C-4´), 151.37 (C-2), 152.04 (C-3)
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a solution of water (25 mL) and glacial acetic acid (5 mL) on
heating in a water bath. Mercury(II) acetate (10.2 g) was dis-
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addition of thioacetamide (2.4 g) and heating in a water bath.
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small volumes of water and then with ethanol. The ethanol fil-
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carbonate.The alkaline solution was extracted with chloroform
and evaporated to dryness in vacuo after drying. Papaverinol 2
was crystallized from the residue obtained in ethanol. This
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