S. Razi et al.
the Kazlauskas rules, where the R-enantiomer reacts faster
than the S-enantiomer during the enzymatic kinetic resolu-
tion of secondary alcohols [28].
water until micro-aqueous media. The reactivity and selec-
tivity of the lipase CAL-B as catalyst was studied on deacyla-
tion of 1-phenylethyl acetate 1a as study model. A series of
experiments of enzymatic deacylation reactions were car-
ried out on 1 mol of rac-phenylethyl acetate (1a) diluted in
2 mL of organic co-solvent in the presence several propor-
tion of phosphate bufer solution (pH 7) with the appropri-
ate amount of lipase. The reaction mixtures were stirred for
48 h at room temperature. Both enantiomers of the remained
esters and furnished alcohols were recovered after fltration
of the lipase and liquid–liquid extraction. The course and
selectivity of the enzymatic hydrolysis were quantifed by
chiral chromatography, and the results are summarized in
acyl chain side (ee>99%), but reactivities varies between
good to high depending of their bulkiness (31≤conv≤50).
The deacylation of 1-phenylethyl butyrate 3a was achieved
with conv = 31% compared to the 1-phenylethyl propion-
ate 2a and the 1-phenylethyl acetate 1a (entry 4 vs entries
3–2). Good results were obtained for the deacylation of fatty
esters 5a–6a with conversion varied 44.7<conv<48.2 and
excellent selectivity E>200 (entries 5–6). Whilst, using a
benzoate as acyl side 7a, the conversion and the selectivity
drops sharply in the three tested solvents, the best, conv=9%
and E = 12 was noted in TBME (Entry 8 vs Entries 7, 9).
This result suggests that the deleterious impact on the con-
formational changes on lipase due to a aryl group impact
more strongly than the efect of chain portion with isopropyl
The results from Table 2 indicate the high enantioselectiv-
ity of the lipase CAL-B under all the experimental conditions
conditions for the conventional hydrolysis [30], an impor-
tant infuence of CAL-B loading on the lipase reactivity is
observed. The frst test is carried out in an ethyl ether/water
mixture (2 mL/12 mL) and with 150 mg of CAL-B (entry
1). More eco-friendly reaction conditions [31–33] were
phate bufer solution also the CAL-B amount with the use
of tertiobutylmethyl ether (TBME) as alternative to of the
diethylether in term of environmental exigencies [34, 35].
the conversion value of 50%, the (R)-alcohols are obtained
with excellent enantiomeric excesses, ees = eep = 99% and
E>200 (entries 1 and 2). No disturb on both reactivity and
selectivity was recorded by more reducing the amount of
CAL-B to lower values of 12.5 mg per 1 mmole of substrate
(entries 6–7).The decreasing of the proportion of the bufer
solution/co-solvent (v/v) from (5/1) to (0.02/1) only a slight
drop of the conversion from 50 to 45% (entry 5 vs 9). No
reactivity without external source of water was observed
(entry 10). So, to ensure the hydrolysis reaction, it’s suf-
cient to use micro-aqueous system under the optimum elabo-
rated conditions and a single drop of water was essential to
catalytic activity. To study the scope of the CAL-B catalyzed
hydrolysis in micro-aqueous media with the lipase CAL B,
we extended the methodology using our optimized condi-
tions to other substrates. These experimental conditions have
been performed on deacylation of various rac-α-phenyl ethyl
esters [1a–7a] in micro-aqueous media with the lipase CAL
B. The obtained results have been compared to those given
The obtained results are detailed in Table 3.
The obtained results are in according of previous results
of enzymatic hydrolysis in biphasic medium, which is
explained by the shape of the active site of the lipase CAL-
B. It is more adapted to accept linear substituent, such as
In the previous investigation, the basic non-conventional
deacylation catalyzed by CAL-B on the aryl ethyl acetates,
had not shown signifcant efect of the aryl moiety, on both
the reactivity and the selectivity, while here the structure
of the acyl side moiety have a signifcant efect on catalytic
performance of the lipase CAL-B.
The aryl substituent as a leaving group shows a drastic
impact on the reactivity and the selectivity of the CAL-B dur-
ing the deacylation with sodium carbonate. It is not the case
with the aliphatic substituent’s, which seem the best candi-
dates for this pathway. This easy non-conventional meth-
odology, assisted by sodium carbonates has been applied
without any external water addition shows ideal for the
cleavage of the aliphatic chain acyl moieties, especially for
the deacylation of fatty esters with high enantioselectivities.
3.2 CAL‑B Catalyzed Kinetic Resolution
of rac‑α‑phenyl Ethyl Esters: From Biphasic
to Micro‑aqueous Media
The objective of this study was to examine the enzymatic
hydrolysis operated in micro-aqueous media. Traditional
enzymatic hydrolysis of racemic esters was mostly operated
within a large amount of external water (phosphate bufer
solution pH 7). To fnd the optimum conditions, the catalytic
amount of the biocatalysts, was frst examined. Then, the
enzymatic hydrolysis have realized under diferent condi-
tions of aqueous medium, from large amount of external
media is carried out with an optimal amount of lipase CAL-
B, the conditions of 12.5 mg of lipase CAL-B in TBME/
H2O (1/0.02) have been selected for the kinetic resolution of
rac-α- phenyl alkyl esters [1a–7a] in micro-aqueous media.
1 3