.
Angewandte
Communications
additional follow-up products were formed, which were
identified as the four stereo- and regioisomeric products of
a spontaneous Pictet–Spengler reaction of 2a with acetalde-
hyde,[18] a side product of the dealkylation. The formation of
acetaldehyde was verified by GC and supported by a Purpald-
based colorimetric assay (see the Supporting Information).
From these data, we conclude that BBE catalyzes oxidative
dealkylation.
To prevent the formation of the side products, an
alternative substrate 1b, which lacks the phenolic alcohol
moiety essential for the spontaneous Pictet–Spengler reac-
tion, was tested for N-dealkylation. Indeed, the transforma-
tion of 1b led exclusively to the dealkylated product 2b.
In a next step, the reaction conditions were optimized.
The most suitable buffer was found to be 50 mm tris(hydrox-
ymethyl)aminomethane hydrochloride (Tris-HCl) at pH 9
and containing 10 mm MgCl2. The hydrogen peroxide formed
as a byproduct was disproportionated to water and molecular
oxygen by a catalase. Reactions were performed in the dark
and the temperature was set to 408C. Since the non-natural
substrates were barely soluble in buffer, various water-
miscible and water-immiscible organic solvents (10% v/v)
were tested for substrate solubilization. In general DMSO
(10% v/v) turned out to be preferred at a substrate concen-
tration of 10 mm.
the methoxy group. In contrast, N-ethyl substituted deriva-
tives bearing a chlorine atom at the para position of the
benzyl group (rac-1d) or a methylenedioxy bridge at the
isoquinoline core (rac-1e) were readily accepted. Both
substrates were dealkylated with excellent stereoselectivity
(E > 100), leading to optically pure secondary amines (S)-2d
and (S)-2e (ee > 98%; Table 1). Again, exclusively the (S)-
enantiomer was transformed. These results also indicate that
both electron-donating and electron-withdrawing substitu-
ents in the para position of the benzyl moiety in substrate
1 are readily accepted.
For comparison, related substrates possessing a N-methyl
instead of the N-ethyl group were tested, although the
compounds orientaline and isoorientaline (Figure 1) had
Under optimized conditions, substrate rac-1b (10 mm) was
transformed into 2b, with 45% conversion within 24 h.[19]
Analysis of the optical purity revealed that the ee of the
remaining substrate (1b) was 74% in favor of the (R)-
enantiomer and that the obtained product (S)-2b was
optically pure (ee > 98%). This corresponds to a kinetic
resolution with an enantioselectivity of E > 100 (Table 1). Out
of the racemic mixture, exclusively the (S)-enantiomer was
transformed, while the (R)-isomer remained untouched.
Substrate rac-1c, which bears the methoxy substituent in
the meta position of the benzyl group, led to clear conversion
into the dealkylated product 2c, although it was converted
slowly, most likely due to steric hindrance in the active site by
Figure 1. The 1-Benzyl-N-methyl-1,2,3,4-tetrahydroisoquinolines (3)
investigated in the present study.
already been reported not to be accepted by BBE.[20]
Laudanosine, which bears two methoxy substituents on the
isoquinoline core, was previously shown to be oxidized slowly
À
at the C1 N bond to give a double bond in conjugation to the
aromatic ring system, but no dealkylation was detected.[14a]
Only for (S)-N-methylcoclaurine, 2% dealkylation was
reported within 6 h at 10 mm substrate concentration.[20]
Consequently, it was unexpected that rac-3a, the N-methyl
homologue to 1b, was readily converted and enantioselec-
tively dealkylated. Within 24 h, 47% conversion was reached,
giving (S)-2b in optically pure form (ee > 98%) and the
remaining substrate (R)-3a in 87% ee (E > 100).
Table 1: Enantioselective N-dealkylation with BBE.[a]
Substrate
c[b] [%]
ee (1)[c] [%]
ee (2)[c] [%]
E[d]
rac-1b[e]
rac-1c
45
4
30
28
46
42
74 (R)
n.d.[h]
41 (R)
21 (R)
80 (R)
63 (R)
>98 (S)
n.d.[h]
>98 (S)
>98 (S)
>98 (S)
>98 (S)
>100
n.a.[i]
>100
>100
>100
>100
However, for other N-methyl compounds such as rac-3b
or rac-3c, no dealkylation was detected, which is in line with
the previously reported N-methyl nonsubstrates.
The substrates tested (1a–e) and product 2e have never
been described before in either optically pure or racemic
form; compounds 3a, 3b, 3c, 2a, 2b, 2c, and 2d have been
described previously but in racemic form only.[16c] Thus,
biocatalytic N-dealkylation with BBE provided access to
novel benzylisoquinoline alkaloid derivatives in optically
enriched or even optically pure form.
Finally, the enantioselective dealkylation was performed
on a preparative scale for substrates 1b and 1e (171 and
81 mg, respectively). After 48 h, the products of the kinetic
resolution (S)-2b and (S)-2e were isolated with good yield
and excellent optical purity (ee > 98%; Table 2).
rac-1d[f]
rac-1d
rac-1e[f]
rac-1e[g]
[a] Reaction conditions: substrate rac-1 (10 mm), Tris-HCl buffer
(50 mm, containing 10 mm MgCl2), pH 9, 10% v/v DMSO, crude
catalase preparation (5 mgmLÀ1), BBE (5 mgmLÀ1), glass vials (4 mL) in
vertical position while shaking, reactions performed in the dark, 408C,
24 h. [b] Conversion was measured by HPLC on a C18 stationary phase.
[c] Enantiomeric excess was measured by HPLC on a chiral stationary
phase. [d] E value calculated from the ee of the substrate and product by
enantio.pl. [e] Glass vials in horizontal positon while shaking. [f] Sub-
strate concentration 5 mm. [g] Substrate concentration 5 mm, enzyme
concentration 3 mgmLÀ1, 10% v/v DMF. [h] Not determined due to low
conversion. [i] Not applicable.
ꢀ 2015 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Angew. Chem. Int. Ed. 2015, 54, 15051 –15054