Epimerization of lupinine to epilupinine and vice versa. Reexamination of the structures of lupinal and epilupinal

Article abstract of DOI:10.1002/hlca.200590005

Although the epimerization of lupinine (1) has been largely investigated, a previously not observed compound of formula C₁₀H₁₇NO was now isolated from the mixture of alkaloids that remains after the separation of epilupinine (2). It is insoluble in dry Et₂O but soluble in EtOH, from which it is recovered as an Et₂O-soluble oil that slowly returns to the Et₂O-insoluble solid form. For these characteristics and based on GC/MS, ¹H-NMR, and IR data, it is considered as the inner salt 6 of the common enolic form 5 of lupinal (3) and epilupinal (4), with which it is in equilibrium when standing in solution (see Scheme 1). The oily form, but not the solid one, is able to improve the conversion of 1 to 2, establishing the role of the aldehydes in the epimerization process. It was observed that also 2 can be converted to 1. Finally, the solid lupinal described by Zaboev should be considered as being identical to the now isolated inner salt 6, while the oily epilupinal of Wicky and Schumann is, indeed, a mixture of epilupinal (4) with a minor amount of lupinal (3), which, on standing, is converted to the inner salt 6 of the common enolic form 5.

Full text of DOI:10.1002/hlca.200590005

Helvetica Chimica Acta ± Vol. 88 (2005)
245
Epimerization of Lupinine to Epilupinine and vice versa. Reexamination of
the Structures of Lupinal and Epilupinal
by Anna Sparatore*a), Bruno Tassob), Vito Boidob), and Fabio Sparatoreb)
a
Á
) Istituto di Chimica Farmaceutica, Universita di Milano, Viale Abruzzi 42, I-20131 Milano
(tel.: ‡ 390250317554; fax: ‡ 390250317565; e-mail: anna.sparatore@unimi.it)
b
Á
) Dipartimento di Scienze Farmaceutiche, Universita di Genova, Viale Benedetto XV, 3, I-16132 Genova
Although the epimerization of lupinine (1) has been largely investigated, a previously not observed
compound of formula C10H17NO was now isolated from the mixture of alkaloids that remains after the
separation of epilupinine (2). It is insoluble in dry Et2O but soluble in EtOH, from which it is recovered as an
Et2O-soluble oil that slowly returns to the Et2O-insoluble solid form. For these characteristics and based on GC/
MS, 1H-NMR, and IR data, it is considered as the inner salt 6 of the common enolic form 5 of lupinal (3) and
epilupinal (4), with which it is in equilibrium when standing in solution (see Scheme 1). The oily form, but not
the solid one, is able to improve the conversion of 1 to 2, establishing the role of the aldehydes in the
epimerization process. It was observed that also 2 can be converted to 1. Finally, the solid lupinal described by
Zaboev should be considered as being identical to the now isolated inner salt 6, while the oily epilupinal of
Wicky and Schumann is, indeed, a mixture of epilupinal (4) with a minor amount of lupinal (3), which, on
standing, is converted to the inner salt 6 of the common enolic form 5.
Introduction. ± For a long time, we have been using (À)-lupinine (1) and (‡)-
epilupinine (2) as the starting material for the preparation of octahydro-2 H-
quinolizine (quinolizidine) derivatives of pharmacological interest.
While lupinine (1) has been extracted from seeds of either cultivated bitter Lupinus
luteus L. or wild Sardinian Lupinus hispanicus Boiss et Reut. [1], epilupinine (2) has
been obtained through epimerization of 1.
The epimerization of 1 to 2 has been accomplished by several authors by heating its
benzene solution for 3 d in the presence of Na [2 ± 5]. Having repeated several times
this experiment, Sparatore and Boido [6] observed that the yield of 2 was largely
variable in an unaccountable way. This observation conformed to that of Clemo and
Rudinger [3], but was in contrast to that of Galinovski and Nesvadba [4].
Better results in the conversion of lupinine (1) to epilupinine (2) were obtained by
Boido and Sparatore [7] by refluxing the xylene solution of 1 for 5 h in the presence of
either NaH, NaNH2 or NaOEt, followed by the isolation of the main portion of 2 and
ꢀ 2005 Verlag Helvetica Chimica Acta AG, Zürich

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repeating twice the same treatment with the residual mixture 1/2; a conversion higher
than 90% was finally obtained.
A reasonable mechanism for the epimerization was reported in 1972 by
Mnatsakanyan et al. [8], who postulated the initial conversion of lupinine (1) to the
corresponding aldehyde, lupinal (3); the latter would then be tranformed, via the enol
form 5, to epilupinal (4), which would finally be reduced to epilupinine (2) (cf.
Scheme 1). All steps are considered unidirectional.
Scheme 1
In the present communication, we illustrate experimental results that substantiate
the proposed mechanism, establishing, however, that it is bidirectional since also
epilupinine (2) can be converted to lupinine (1). The structures of compounds
previously described as lupinal and epilupinal are also reexamined.
Isolation of (Quinolizidin-1-ylidene)methanol Inner Salt (6). ± In an attempt to
recover some more epilupinine (2) from the residues of the many epimerization
experiments with lupinine (1) performed so far, we isolated a new compound 6, which
throws some light on the epimerization process 1 ! 2. Compound 6 (m.p. 100.5 ± 1038)
is very poorly soluble in dry Et2O and its elemental analysis revealed a formula
C10H18NO1.5 corresponding to the hemihydrate of a compound with two H-atoms fewer
than 1 or 2 (C10H17NO ´ 0.5 H2O). Therefore, 6 could be considered a didehydro
derivative of either 1 or 2 but is different from lupinal (3; m.p. 93 ± 968) [9] and
epilupinal (4; oil) [10]. The molecular formula of 6 was confirmed by MS, which
exhibited a molecular-ion peak at m/z 167, i.e., at two mass units lower than that of 1
1
and 2 (m/z 169). The H-NMR spectrum (CDCl3) of 6 suggested the presence of an
enol besides traces of an aldehyde.

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The MS of 6 was largely coincident with the spectra of (1) and epilupinine (2) [11], which, in turn, differ
from each other only slightly in the relative abundance of fragment-ion peaks. Nevertheless, two main
‡
differences should be pointed out. The relative intensity of the M of 6 was very low (3%), while that of 1 and 2
is very high (60and 90%, resp.). Moreover, the fragment at m/z 152 is the base peak (100%) of 1 and 2 arising
from the loss of the OH group, whereas, very significantly, this fragment ion was lacking in the MS of 6. Instead,
a peak at m/z 150, corresponding to the loss of an OH group, was present with a very low abundance (4%),
suggesting a quite different molecular entourage in 6 from that of the enol form of the aldehyde.
The 1H-NMR spectrum of 6 exhibited a d at d 9.59 (J ˆ 3.5 Hz), that might be related to the proton of an
aldehyde group coupling with HÀC(1) of the quinolizidine ring. However, the integration of the d accounted for
only 0.14 ± 0.31 H, depending on the concentration of the CDCl3 solution. The low intensity of this peak could be
due either to an equilibrium between the carbonyl and the enol forms or by a reversible interaction of the
solvent with the carbonyl group. The presence of exchangeable protons around d 1.9 (superimposed to the
quinolizidine protons) might support the presence of an enol OH group, though no quantitative evaluation
could be performed due to the presence of H2O, which could not be removed completely from the hemihydrate.
The IR spectrum (KBr) of 6 exhibited a low-intensity carbonyl band at 1722 cmÀ1
and a strong broad band around 3260cm À1, supporting further the existence of an
equilibrium between an aldehyde and its enol form, either as such or as a salt with the
strongly basic quinolizidine N-atom. The formation of a salt would account for the very
poor solubility in dry Et2O.
Compound 6 was very soluble in EtOH, but the oily residue obtained upon
evaporation of the EtOH solution was now also soluble in dry Et2O. After evaporation
1
of the Et2O solution, the oily residue exhibited the same H-NMR and IR spectra as
those already discussed for the Et2O-insoluble compound, but the intensities of the
aldehyde signals (d 9.6; 1722 cmÀ1) were enhanced, while those of the enol signals (d
1.9; 3260cm À1) were reduced. On standing, the oily compound returned slowly to the
Et2O-insoluble solid form.
Finally, reduction of 6 with NaBH4 gave rise to a mixture of lupinine (1) and
epilupinine (2); 1 representing a quite minor component of the mixture, as indicated by
the integration of its peculiar 1H-NMR signal at d 4.17 (ddd, J ˆ 10.4 and 4.6 Hz) [12],
due to the long-range coupling of one of the protons of the exocyclic methylene group
with the OH proton bound to the tertiary N-atom, as depicted in structure 1.
All these observations allowed us to attribute to the novel compound 6 the structure
of the inner salt of (quinolizidine 1-ylidene)methanol, which is the common enol form
of both lupinal (1) and epilupinal (2) and, with which 6 is in a tautomer equilibrium
when it is dissolved in EtOH or CDCl3.
Mechanism of Epimerization of Lupinine (1) and Epilupinine (2). ± The always
incomplete conversion of lupinine (1) to epilupinine (2) suggests the presence of a
reversible process tending to an equilibrium. To substantiate this assumption, we
heated separately for 6 h at 1658 in closed tubes very pure 1 and 2 in xylene solution in
the presence of Na. In both cases, we obtained a mixture 1/2, thus establishing the
reversibility of the epimerization process. The conversion 1 ! 2 was ca. 30%, while 2 !
1 was 11 ± 14% (result of several experiments). The addition of a small amount of
(quinolizidin-1-ylidene)methanol inner salt (6) did not improve the conversion 1 ! 2;
however the addition of the oily form of 6 (see above) raised the yield of 2 to 67%.
Extending the heating time to 9 h, the yield increased to 85%, but further heating did
not improve it anymore .The conversion 2 ! 1 was not affected by the addition of the
oily form of 6.

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Therefore, the reciprocal epimerization of 1 and 2 may be represented by Scheme 1.
Lupinal (3), required to initiate the epimerization of 1, could be formed either by air
oxidation or, better, by thermal elimination of NaH from sodium lupinine (1a); then
epilupinal (4) by reacting with sodium lupinine (1a) forms sodium epilupinine (2a) and
a new amount of lupinal (3) that will propagate the reaction (Scheme 2). Starting from
epilupinine (2) the sequence will proceed in the reverse direction. The formation of the
insoluble inner salt 6, by subtracting the aldehydes from the system, is detrimental on
the epimerization process which can proceed only if new lupinal is produced.
Scheme 2
Reexamination of the Structures of Lupinal (3) and Epilupinal (4). ± So-called
lupinal was obtained by Zaboev [9] by oxidation of lupinine (1) with CrO3 in AcOH
and was described as an Et2O-insoluble compound melting at 93 ± 968, only a few
degrees lower than our inner salt 6 of (quinolizidine-1-ylidene)methanol. On the other
hand, so-called epilupinal was obtained by Wicky and Schumann [10] on oxidation of
epilupinine (2) with DMSO in the presence of dicyclohexylcarbodiimide and
anhydrous phosphoric acid according to the method of Pfitzner and Moffatt [13]. This
epilupinal was described as an oil, and, for its characterization, the 2,4-dinitro-
phenylhydrazone was prepared (probably by the use of a sulfuric acid solution of the
hydrazine); however, the elemental analysis of this derivative did not account for the
proposed structure. If the given values for H and N are exchanged, the analytical results
agree with those required for the hydrogen sulfate of this epilupinal 2,4-dinitrophe-
nylhydrazone. Nevertheless, the MS of this oily aldehyde is in reasonable accordance
with that of our ether-insoluble compound 6.
To clarify the situation, we oxidized lupinine (1) and epilupinine (2) with DMSO
following the procedure of Wicky and Schumann [10]. In both cases, identical oily
‡
mixtures of the two aldehydes 3 and 4 with an M at m/z 167 were obtained, as
established by GC/MS (Scheme 3). The two aldehydes, differing in retention times by
only 0.57 min, were present in a ratio ranging from 5 :1 to 9 :1 in repeated experiments.
The fragmentation patterns of the two aldehydes were identical and were also identical
to those of the product isolated from the epimerization mixtures. The initially oily
mixture obtained from DMSO oxidation of 1 and 2 exhibited a clear carbonyl band in
the IR spectrum, but, on standing, the mixtures solidified slowly with concomitant
decrease in the intensity of the carbonyl band; finally they became completely solid and
Et2O-insoluble. Since identical aldehyde mixtures were obtained from 1 and 2, the

Helvetica Chimica Acta ± Vol. 88 (2005)
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interconversion of the two epimeric aldehydes to reach an equilibrium must be very
easy. The NaBH4 reduction of the initially oily aldehyde mixtures gave mixtures of
lupinine (1) and epilupinine (2), with the latter largely prevailing over the former.
Scheme 3
Therefore, it seems sound to conclude that the solid so-called lupinal obtained by
Zaboev is, indeed, the inner salt 6 of the enol form 5, while the oily so-called epilupinal
of Wicky and Schumann is a mixture of the two epimeric aldehydes, epilupinal (4) and
lupinal (3), not yet converted to the inner salt 6 of the common enol form 5.
It is worth noting that the oxidation of 1 and 2 with DMSO was accompanied by the
formation of an S-containing compound of molecular mass 229 that might be the
(methylthio)methyl ether of the starting alcohol (see Scheme 3). Indeed, traces of such
(methylthio)methylethers of the starting alcohol have been already isolated by Pfitzner
and Moffatt [13], who also proposed the mechanism for their formation. In our case,
particularly from 1, the supposed (methylthio)methyl ethers were formed in substantial
quantities, accounting for the relatively low yield of the aldehydes.
Experimental Part
General. All commercially available solvents and reagents were used without further purification, unless
otherwise stated. TLC: aluminium oxide 60F254, neutral (Merck). M.p.: Büchi apparatus; uncorrected. Optical
activity; Perkin-Elmer 241 MC polarimeter; sodium lamp, l 589 nm; tube length 10cm; EtOH solns. IR Spectra:
Perkin-Elmer Paragon-1000-PC spectrophotometer; KBr pellets for solids, and neat for liquids; nÄ in cmÀ1
.
1H-NMR Spectra: Varian Gemini-200 spectrometer; in CDCl3 with Me4Si as internal standard; d in ppm, J in
Hz. GC/MS: Hewlett-Packard 6890/5973 equipment; m/z (rel.%). Elemental analyses were performed on a
Carlo Erba EA-1100 CHNS-O instrument in the Microanalysis Laboratory of the Department of Pharmaceut-
ical Sciences of Genoa University.
Lupinine Epimerization: (Hexahydro-2H-quinolizin-1(6H)-ylidene)methanol Inner Salt (6). To a soln. of
(À)-lupinine (1; 8 g) in dry xylene (40ml), NaH (3 g of 60% dispersion in mineral oil) was added, and the
mixture was heated under reflux for 5 h under stirring (e.m.). On reaching the boiling temp., the mixture
became pasty. After further stirring overnight at r.t., the mixture was extracted with 6n HCl (25 ml) and 0.3n
HCl (2 Â 25 ml), and then with H2O. The acid soln. was extracted twice with Et2O, strongly basified with 30%
KOH soln., and extracted with Et2O. The aq. phase was saturated with K2CO3 and further extracted with Et2O.
The Et2O soln. was dried (Na2SO4) and evaporated. The obtained oil (7.6 g) was dissolved in dry Et2O (5 ml)
and left in the cold. Crystals were collected and washed with cold dry Et2O/light petroleum ether 3 :7. On

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concentration of the Et2O soln., a second crop of crystals was collected and washed as above: 4.56 g (57%) of
(‡)-epilupinine (2). M.p. 74.4 ± 75.48.
After evaporation of the remaining Et2O soln., 3 g of oily crystals were recovered, which (eventually joined
with analogous fractions obtained from other epimerization experiments) were treated again with NaH as
above. The whole procedure might be repeated improving the yield of 2. The further crops of crystals of 2
melted at 72 ± 788 with a few crystals melting around 908; these portions of 2 were dissolved in dry Et2O leaving a
whitish residue (m.p. 100 ± 1038, see below), while from the evaporated soln., purer 2 was recovered. M.p. 77 ±
788 (petroleum ether) [a]2D0 ˆ ‡34.53 (958/EtOH, c ˆ 0,94).
The whitish, Et2O-insoluble (ca. 200 mg from each batch (8 g) of 1 used; i.e., 2.5% yield), was boiled
several times with dry Et2O to eliminate any residual 1 and 2: pure 61) M.p. 100.5 ± 1038. IR (KBr): 1722 (CˆO),
‡
1
3150, 3250 (OH and/or NH ). H-NMR (CDCl3): 1.1 ± 2.2 (m with superimposed s at 1.9, ca. 13.7 H, 13 H of
quin. ca. 0.7 H of OH collapsing with D2O); 2.7 ± 2.9 (m, 2 H, a to N); 3.50± 3.74 ( m, 1 H of quin.); 9.59 (d, J ˆ
‡
‡
‡
‡
3.5, ca. 0.3 H, CHˆO). MS: 167 (3, M ), 166 (12, M À H] ), 150(4, [ M À OH] ), 138 (62, [M À CHˆO] ), 124
(10), 110 (42), 96 (51), 83 (100), 67 (3), 55 (11). [a]2D0 ˆ ‡10.08 (95% EtOH, c ˆ 0.594). Anal. calc. for
C10H17NO ´ 0.5 H2O: C 68.14, H 10.29, N 7.95; found: C 68.26, H 9.94, N 8.16.
Reduction of Compound 6. To a soln. of 6 (150mg) in 80% EtOH (5 ml), NaBH 4 (50mg) was added. The
soln. was refluxed for 3 h and then evaporated. The residue was taken up in H2O, treated with a few drops of 6n
NaOH and extracted with Et2O. The org. phase was dried (Na2SO4) and evaporated. 135 mg of 1/2 as oily
crystals. TLC (alumina, iPrOH): Rf 0.32 (1; minor) and 0.44 (2). 1H-NMR (CDCl3): 4.10± 4.25 ( m, ca. 0.15 H)
indicating a 15% content of 1; for pure 1 this signal is at 4.17 (ddd, J ˆ 10.4 and 4.6); it is absent in the spectrum
of 2.
Reciprocal Conversion of Lupinine (1) and Epilupinine (2). Pure 1 (500 mg, m.p. 69 ± 708, [a]2D0 ˆ À21.498)
was dissolved in dry xylene (3 ml) in an Aldrich pressure tube. Na (70mg) was added, and the closed tube was
heated in an oil bath at 1658 for 6 h. The metal slowly disappeared, while a viscous mass was formed. After
cooling, the mixture was diluted with Et2O and extracted several times with dil. HCl soln. The acid soln. was
alkalinized with 6n NaOH, saturated with NaCl, and extracted with Et2O. The org. phase was dried (Na2SO4)
and evaporated, leaving 478 mg of an oil of. [a]2D0 ˆ À5.00 (95% EtOH, c ˆ 0.956), corresponding to a mixture of
70.6% of 1 and 29.4% of 2.
Repeating the same experiment (470mg of 1) in the presence of the solid 6 (30mg), the degree of
conversion of 1 to 2 was practically the same.
In a further experiment, with 30mg of the oily compound obtained from the EtOH soln. of 6 or from
oxidation of 1 (see below), 464 mg of an oil of [a]2D0 ˆ ‡16.04, were obtained, corresponding to a mixture of
33% of 1 and 67% of 2. Extending the heating time to 9 h, the yield raised to 85%.
Starting from pure 2 (500 mg; m.p. 77 ± 788, [a]D20 ˆ ‡34.53) and operating under the same conditions,
456 mg of an oil of [a]2D0 ˆ ‡26.85 (95% EtOH, c ˆ 0.912) were obtained that soon crystallized, corresponding to
a mixture of 13.7% of 1 and 86.3% of 2. Also in the presence of the oily aldehyde, the yield of 1 remained at ca.
15%.
The optical activity of mixtures of 1 and 2 changed linearly with composition.
Oxidation of Lupinine (1). Pure 1 (1 g, 5.9 mmol) was dissolved in anh. DMSO (40ml; distilled in vacuo
and stored over 4-Š molecular sieves) containing dicyclohexylcarbodiimide (3.2 g, 16 mmol). Anh. H3PO4
(1.2 g, 12 mmol) was added, and the mixture was stirred at r.t. for 1 h. Under ice cooling, 2n NaOH (20ml) was
added, and the white precipitate was centrifugated. The precipitate and the aq. DMSO soln. were extracted with
petroleum ether. The combined org. phase was filtered and evaporated and the oily residue taken up in dil. HCl
soln. and washed with Et2O. The acid soln. was basified and extracted with Et2O, the extract evaporated, and the
residue (0.76 g) bulb-to-bulb distilled at 0.3 Torr. At 110 ± 1158 (air-bath temp.), 0.54 g of colorless oil were
‡
collected. GC/MS: two aldehydes with M at m/z 167 (tR 15.1 and 15.6; ratio 7:1, varying to up to 9 :1 in further
‡
experiments), 1 with M at m/z 169 (tR 16.4), and traces of the supposed O-[(methylthio)methyl]lupinine with
‡
M
at m/z 229 (tR 20.0), the latter being the main component of the distillation tail. On standing, the distilled oil
solidified, and after repeated washings with dry Et2O, a white powder (230mg, ca. 23% yield; m.p. 95 ± 968) was
‡
obtained, which exhibited a unique peak in the GC/MS: M at m/z 167. The MS fragmentation patterns of both
oily and solid aldehydes were identical with that of the compound isolated from the epimerization mixture. IR
‡
(KBr): 3260s (OH and/or NH ), 1722w (CˆO).
1
)
This compound is very soluble in EtOH, from which it is recovered as on oil with practically the same
spectral characteristics as described above.

Helvetica Chimica Acta ± Vol. 88 (2005)
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Oxidation of Epilupinine (2). Pure 2 was oxidized as described for the oxidation of 1 with practically the
same results, apart from a higher yield (40± 50%) of solid aldehyde. Also in this case, the GC/MS of the initially
‡
oily aldehyde indicated the presence of two fractions with M 167 in a ratio 7 ± 9 :1.
Reduction of Prepared Aldehydes. The NaBH4 reduction of either the oily or solid aldehydes obtained from
both 1 and 2 gave always similar mixtures of 1 and 2 (TLC), with a large prevalence of the latter, similar to the
reduction of 6 isolated from the epimerization mixture.
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Received October 25, 2004