Lipase catalysed kinetic resolution of racemic 1,2-diols containing a chiral quaternary center

Article abstract of DOI:10.3390/molecules23071585

Optically active 1,2-diols are valuable buildings blocks in organic synthesis. In the present paper, a set of racemic 1,2-diols with an ester functional group are prepared, starting from α-ketoesters in a three-step procedure with moderate yields. The racemic 1,2-diols, containing a chiral quaternary center in their structure, are subjected to selective acylation in order to perform their kinetic resolution catalysed by a set of commercially available lipases. Under optimized reaction conditions, good conversions and enantioselectivities are achieved by using the lipase PSL-C from Pseudomonas cepacia in tert-butyl methyl ether. This biocatalyst could be reused up to five times without losing its properties.

Full text of DOI:10.3390/molecules23071585

molecules
Article
Lipase Catalysed Kinetic Resolution of Racemic
1,2-Diols Containing a Chiral Quaternary Center
ID
Gonzalo de Gonzalo
Departamento de Química Orgánica, Universidad de Sevilla, c/Profesor García González 1, 41012 Sevilla, Spain;
gdegonzalo@us.es; Tel.: +34-954559997
ꢀꢁꢂꢀꢃꢄꢅꢆꢇ
ꢀꢁꢂꢃꢄꢅꢆ
Received: 25 May 2018; Accepted: 27 June 2018; Published: 29 June 2018
Abstract: Optically active 1,2-diols are valuable buildings blocks in organic synthesis. In the present
paper, a set of racemic 1,2-diols with an ester functional group are prepared, starting from α-ketoesters
in a three-step procedure with moderate yields. The racemic 1,2-diols, containing a chiral quaternary
center in their structure, are subjected to selective acylation in order to perform their kinetic resolution
catalysed by a set of commercially available lipases. Under optimized reaction conditions, good
conversions and enantioselectivities are achieved by using the lipase PSL-C from Pseudomonas
cepacia in tert-butyl methyl ether. This biocatalyst could be reused up to five times without losing
its properties.
Keywords: biocatalysis; lipases; kinetic resolution; 1,2-diols
1. Introduction
Optically active 1,2-diols are valuable compounds as they can be transformed into several
interesting molecules [
of synthetic methodologies have been developed for their preparation [
applied methods is Sharpless asymmetric dihydroxylation, which involves the oxidation of alkenes
to form 1,2-diols in presence of chiral catalysts [ ]. This methodology suffers some limitations
1–
3]. Due to their importance as building blocks in organic synthesis, a number
4,5]. One of the most widely
6,7
including the relatively low activity and selectivity of the aromatic compounds and the substrate
scope. For these reasons, different alternatives have been investigated. The use of biologically active
systems as catalysts in organic reactions, including whole cells, cells free extracts, or purified enzymes,
has emerged as a powerful tool for the preparation of high added value compounds under mild and
environmentally friendly conditions [8–11].
The enzymes that have been used to catalyse the formation of chiral 1,2-diols belong to the
oxidoreductases (alcohol dehydrogenases and dioxygenases) [12
epoxide hydrolases [14 15] and lipases. This last group of enzymes (EC 3.1.1.3) have demonstrated
their synthetic applicability, being the most used type of enzymes in industrial chemistry [16 18].
,13], and the hydrolases, including
,
–
Lipases are widely available, have broad substrate acceptance and are able to catalyse reactions not
only in aqueous mediums, but also in organic solvents, which expands their synthetic repertoire.
In addition, lipases usually display a high degree of chemo-, regio- and/or enantioselectivity in the
processes that they catalyse.
Lipases have been used to prepare optically active 1,2-diols catalysing the kinetic resolution of
racemic mixtures in acylation reactions, thus leading to chiral diols and esters that can be converted
back to the starting diols via hydrolysis [19–22]. In 2000, the kinetic resolution of racemic ethyl
2-benzyl-2,3-dihydroxypropanoate was described, a 1,2-diol containing an ester moiety precursor in
the synthesis of (R)-etomoxir, which is a powerful hypoglucemic reagent [23]. After testing different
biocatalysts, the lipase PS from Amano was found to catalyse the formation of (R)-1,2-diol and
(S)-acetate with high selectivity and activity. Given this result, we decided to synthesise a set of
Molecules 2018, 23, 1585; doi:10.3390/molecules23071585
www.mdpi.com/journal/molecules

Molecules 2018, 23, 1585
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functionalised 1,2-diols with an ester group and perform their kinetic resolution in the presence of
different lipases, with the aim of obtaining these valuable optically active compounds.
2. Results and Discussion
2.1. Preparation of the Racemic 1,2-Diols (±)-1–6d
The racemic 1,2-diols were prepared in a three-step procedure starting from the corresponding
α
-ketoesters
1
–
6a, as indicated in Table 1. These compounds were treated with N-tert-butyl
)- 6b
formaldehyde hydrazone in toluene at room temperature to yield the racemic azocompounds (
with high yields (75–92%) after 24 h. For almost all the ketoesters, the reaction occurred in the absence
±
1–
of any catalyst, but for ethyl benzoylformate (1a), the addition was accelerated in the presence of
the Schreiner’s thiourea (
be transformed in different compounds [24
(aq) yielded the corresponding hydroxyaldehydes (
and hydrolysis process. Debt to their instability were directly reduced without purification to the
racemic 1,2-diols ( )- 6d by treatment with a mild reductant as tetrabutylammonium borohydride
I
) at 10 mol%. The resulting azocompounds are valuable synthons that can
26]. Their hydrolysis in a biphasic system Et2O/HCl
)- 6c after four hours via a tautomerization
–
±
1–
±
1–
(NBu4BH4) in CH2Cl2 at room temperature over two hours. Depending on the substrate structure, the
1,2-diols were obtained with yields from 41 to 57%. Attempts to improve these yields by modifying
certain reaction parameters as the hydrolysis conditions or the reducing agent were unsuccessful.
Table 1. Synthesis of racemic 1,2-diols (
commercially available α-ketoesters 1–6a.
±
)-1–6d in a three-step procedure starting from the
Entry
R1
R2
n
Yield (±)-1–6b a
Yield (±)-1–6d a
(±)-1b, b 87
(±)-2b, 80
(±)-3b, 92
(±)-4b, 85
(±)-5b, 90
(±)-6b, 85
b
1
2
3
4
5
6
Ph
Ph
Et
Me
Et
Et
Et
0
0
0
0
0
2
(±)-1d, 47
(±)-2d, 49
(±)-3d, 55
(±)-4d, 45
(±)-5d, 41
(±)-6d, 57
4-CN-Ph
4-OMe-Ph
2-Thiophenyl
H
Et
a
For reaction conditions, see Materials and Methods. Reaction performed with 10 mol% of catalyst I.
2.2. Kinetic Resolution of Racemic Diols (±)-1–6d
Once the racemic 1,2-diols were synthesized, their kinetic resolution was performed. Our initial
studies were performed using racemic ethyl 2,3-dihydroxy-2-phenylpropanoat◦e (
±
)-1d as the model
substrate. The selective acetylation of this 1,2-diol (0.15 M) in toluene at 30 C was performed in

Molecules 2018, 23, 1585
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presence of 3.0 equivalents of vinyl acetate to ensure an irreversible acylation process. The effects of
different biocatalysts were analysed by screening several commercially available lipases, as indicated
in Table 2. For all the biocatalysts tested, the (S)-enantiomer of the 1,2-diol was acetylated, yielding
(S)-1e, whereas the (R)-1,2-diol remained unaltered. The use of immobilized Candida antarctica lipase B
(CalB) resulted in a reaction without selectivity (entry 1), achieving a 23% conversion after four hours.
When the reaction was catalysed by its isozyme A (entry 2), a more selective process was observed
(enantioselectivity, E = 12), demonstrating acylation slower than with CalB. As shown in entry 3,
Pseudomonas cepacia lipase (PSL-C) seems to be the most suitable biocatalyst for this reaction, as a
moderate E value was obtained in a process with a 32% conversion after 20 h, achieving (S)-1d with
86% ee. The use of lipases from Pseudomonas fluorescens (PSF), Burkholderia sp. (BSL), Rhizopus oryzae,
and Aspergillus oryzae led to very low enantioselectivities (E < 10, entries 4–7), with conversions varying
from 41% after 20 h with PSF, to 16% with the same duration using the lipase from Aspergillus oryzae.
The kinetic resolution catalysed by porcine pancreatic lipase (PPL) in toluene afforded a selectivity
value of 13, and a conversion close to 50% after 24 h, as shown in entry 8. The opposite lipase from
Mucor miehei was not a suitable biocatalyst for this reaction, as only a 13% of (S)-1e was obtained after
24 h in a very low selective resolution (entry 9).
Table 2. Lipase-catalysed acylation of rac-ethyl 2,3-dihydroxy-2-phenylpropanoate (1d) at different
reaction conditions.
◦
ee 1d (%) b
E d
Entry
Lipase
Solvent
T ( C)
t (h)
c (%) a
ee 1e (%) c
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
CalB
CalA
PSL-C
PSF
Toluene
Toluene
Toluene
Toluene
Toluene
Toluene
Toluene
Toluene
Toluene
TBME
30
30
30
30
30
30
30
30
20
30
30
30
30
30
30
15
30
30
4
23
32
32
41
38
23
16
48
13
42
41
38
23
6
11
37
40
35
42
22
11
67
12
62
54
50
25
6
37
77
86
50
69
75
57
73
78
91
79
77
86
87
83
91
91
91
2
12
20
20
20
20
20
24
24
12
6
12
20
20
12
24
24
48
12
20
4
8
9
4
13
9
41
17
13
17
15
22
43
37
34
BSL
R. oryzae
A. oryzae
PPL
M. miehei
PSL-C
CalA
TBME
TBME
1,4-Dioxane
THF
PPL
PSL-C
PSL-C
PSL-C
PSL-C
PSL-C e
PSL-C f
DIPE
TBME
TBME
TBME
45
43
36
34
67
67
52
47
a
b
Conversion, c = ees/(ees + eep).
Enantiomeric excesses were determined by high performance liquid
c
chromatography (HPLC) after acetylation in presence of acetic anhydride in pyridine. Determined by HPLC.
d
e
Enantioselectivity (E) value, E = ln[1
−
c(1 + eep)]/ln [1
−
c(1
−
eep)]. Reaction performed with isopropenyl
f
acetate as acyl donor. Reaction performed with ethyl acetate as acyl donor.
After selecting PSL-C as the best biocatalyst for the acetylation of racemic 1d, we analysed other
parameters that can affect the activity and selectivity of the biocatalyst. Thus, different organic solvents
were tested in the acylation reaction. As shown in entry 10, tert-butyl methyl ether (TBME) was the best
solvent for this process, as a good selectivity value could be achieved (E = 41) in a reaction much faster
than in toluene (34% conversion after eight hours with 91% ee for 1e). This solvent was also tested in
the CalA-catalysed acylation, promoting an increase in both the enzyme activity and selectivity (entry

Molecules 2018, 23, 1585
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11). However, the E value remained low. The reaction catalysed by PPL in TBME afforded the chiral
acetate (S)-1e with 77% ee in a process with a 38% conversion after 12 h (entry 12). Other solvents
analysed in the acetylation catalysed by PSL-C, such as 1,4-dioxane or THF, led to slower resolutions
and especially for the latter, whereas the enantioselectivities were around 15. The use of diisopropyl
ether (DIPE) afforded (R)-1d with 67% ee and (S)-1e with 83% ee in a process with a 45% conversion
after 12 h (entry 15).
The effect of temperature was also analysed in this kinetic resolution, performing the
PSL-C-catalysed reaction of (
)-1d at 15 ◦C, as shown in entry 16. Lowering the temperature had no
±
effect on the enzyme selectivity, whereas, as expected, the enzyme activity dropped (c = 25% after 12 h).
The use of a less reactive acyl donor, such as isopropenyl acetate (entry 17), led to a similar selectivity.
Chiral acetate (S)-1e was obtained with a 36% conversion and 91% ee after 16 h. When ethyl acetate
was used as acyl donor (entry 17), a slower kinetic resolution was achieved, as 48 h were required to
obtain a 34% yield of (S)-1e with 91% ee (entry 18).
The recycling of the PSL-C was studied in the selective acetylation of ( )-1e with vinyl acetate
±
in TBME at 30 ◦C. After 20 h, the biocatalyst was filtered, washed with TBME, and used again in a
new reaction cycle. As shown in Figure 1, this biocatalyst could perform the selective acylation of
the racemic diol for five cycles while maintaining its activity and selectivity. For the sixth reaction,
a significant drop in the enantioselectivity of the process was observed (E = 27). For the seventh
reaction, this drop was accompanied by an appreciable loss in enzymatic conversion.
Figure 1. Effect of the PSL-C recycling on the conversion (blue line) and on the enantioselectivity (red
◦
bar) in the catalysed kinetic resolution of (
acetate as he acyl donor.
±
)-1d in tert-butyl methyl ether (TBME) at 30 C using vinyl
After the optimized conditions were set up for the kinetic resoluti◦on of racemic ethyl
2,3-dihydroxy-2-phenylpropanoate, using PSL-C and CalA in TBME at 30 C and vinyl acetate
as acyl donor, the scope of the reaction using different aromatic 1,2-diols was studied (Table 3).
For all the substrates, (S)-acetates
enatioselectivities in all the aromatic and the heteroaromatic substrates (
higher activity. Thus, the enzymatic acylation of racemic methyl 2,3-dihydroxy-2-phenylpropanoate
2d) catalysed by PSL-C occurred with the same selectivity as for the ethyl analogue 1d (E = 42, entry
2
–6e were the obtained products. The use of PSL-C led to higher
2–
5a), whereas CalA showed
(
1) and with a higher conversion, achieving a 41% conversion after 12 h. When the acylation was
catalysed by CalA (entry 2) (S)-2e was obtained with 80% ee and a 42% conversion after eight hours
(E = 16). The p-cyano derivative ( )-3c was a good substrate for both catalysts (entries 3 and 4).
±
When using PSL-C, a 44% of (S)-3e with 90% ee was obtained after 12 h, in a resolution with a good
enantioselectivity value (E = 40), whereas the resolution catalysed by CalA led to a 47% conversion

Molecules 2018, 23, 1585
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after eight hours and a good selectivity (E = 30). The presence of an electron-donating group in the
aromatic ring of the diol seemed to have a negative effect on both the activity (41% of (S)-4e after
24 h) and the selectivity (E = 32) of PSL-C (entry 3). This substrate was tested with CalA, but a low
selectivity was observed (E = 11), in a kinetic resolution with a 36% conversion after 16 h, as shown
in entry 6. A heteroaromatic 1,2-diol as ( )-5d was successfully resolved by PSL-C in MTBE. After
±
24 h, a 45% of (S)-5e with 88% ee was achieved in a process with good selectivity, as shown in entry 7.
The use of CalA led to a 38% conversion after 16 h and moderate selectivity (E = 17, entry 8).
Table 3. PSL-C catalysed kinetic resolution of racemic diols 2–6d in tert-butyl methyl ether (TBME) at
◦
30 C using vinyl acetate as the acyl donor.
ee 2-6d (%) b
E d
Entry Substrate
Lipase
R1
R2
n
t (h)
c (%) a
ee 2-6e (%) c
1
2
3
4
5
6
7
8
9
(±)-2d
(±)-2d
(±)-3d
(±)-3d
(±)-4d
(±)-4d
(±)-5d
(±)-5d
(±)-6d
(±)-6d
(±)-6d
(±)-6d
PSL-C
CalA
PSL-C
CalA
PSL-C
CalA
PSL-C
CalA
Ph
Ph
Me
Me
Et
Et
Et
0
0
0
0
0
0
0
0
2
2
2
2
12
8
12
8
24
16
24
16
2
41
42
44
47
41
36
45
38
45
17
15
17
63
59
71
77
62
42
73
47
50
17
14
18
91
80
90
86
89
72
88
83
62
82
80
86
42
16
40
30
32
11
33
17
7
4-CN-Ph
4-CN-Ph
4-OMe-Ph
4-OMe-Ph
2-Thiophenyl Et
2-Thiophenyl Et
Et
PSL-C
CalA
PSL-C
Ph
Ph
Ph
Ph
Et
Et
Et
Et
10
11 e
12 f
a
6
6
4
12
10
16
PSL-C
b
Conversion, c = ees/(ees + eep). Enantiomeric excesses were determined by HPLC after acetylation in presence of
acetic anhydride in pyridine. Determined by HPLC. d Enantioselectivity (E) value, E = ln[1
c(1 − eep)]. Reaction carried out with ethyl acetate as acyl donor. Reaction carried out at 10 ◦C.
−
c(1 + eep)]/ln [1
−
c
e
f
Regarding the aliphatic diol ( )-6d, in which the stereogenic center presents an aliphatic
±
substituent, the enzymatic acylation in TBME at 30 ◦C using PSL-C afforded a very low
enantioselectivity value (E = 7) in a very fast resolution, achieving a 45% of (S)-6e after 2 h. In view
of this result, the isozyme A from Candida antarctica was tested, leading to a slower (c = 17% after
6 h), but slightly more selective process (E = 12) than with PSL-C (see entry 10). In order to improve
the reaction selectivity, the PSL-C-catalysed resolutions were carried out using ethyl acetate as a less
reactive acyl donor. After 6 h, a 15% of (S)-6e was obtained in a process with a low enantioselectivity
(E = 10). Finally, the kinetic resolution in presence of vinyl acetate was conducted at 10 ◦C. After 4 h,
a 17% of diol 6d was converted into the acetate (S)-6e with 86% ee, but the E value was only increased
to 16 (entry 12), indicating that this substrate was not appropriate for the biocatalysed acylation.
3. Materials and Methods
Unless otherwise noted, analytical grade solvents and commercially available reagents were
used without further purification. Formaldehyde tert-butyl hydrazone [27] and organocatalyst
I
[
28] were synthesized according to the literature. Racemic azocompounds (
±
)-1–6b [24,26] and
1,2-diols ( )-1d and ( )-4,5d [29] exhibited the same physical and spectral properties as described
in the bibliography and the nuclear magnetic resonance (NMR) data of the 1,2-diols are shown
±
±
in the Supplementary Information. Pseudomonas cepacia lipase PSL-C (1638 U/g) and lipases from
Pseudomonas fluorescens (
Burkholderia sp. (≥160 U/mg), porcine pancreatic (
were purchased from Sigma-Aldrich (Saint Louis, MO, USA). Candida antarctica lipase type B (CalB,
≥
160 U/mg), Rhizopus oryzae (
≥
30 U/mg), Aspergillus oryzae (ca. 50 U/mg),
≥
20,000 U/mg), and Mucor miehei (
≥
4000 U/mg)

Molecules 2018, 23, 1585
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Novozyme 435, 7300 propyl laureate units per gram) was obtained from Novozymes (Bagsvaerd,
Denmark). Candida antarctica lipase A (CalA) was obtained from Codexis (Redwood City, CA, USA).
NMR spectra were recorded in CDCl3 [1H-NMR (300 MHz); 13C-NMR (75.4 MHz)] with
1
the solvent peak used as the internal reference (7.26 and 77.0 ppm for H and 13C, respectively).
High-resolution mass spectrometry (HRMS) analyses were performed with an Orbitrap ELITE
instrument (Waltham, ThermoFisher, MA, USA). Column chromatography was performed on silica
gel (Merck Kieselgel 60, Kenilworth, NJ, USA). Analytical thin layer chromatography (TLC) was
performed on aluminum backed plates (1.5
×
5.0 cm) precoated (0.25 mm) with silica gel (Merck,
Silica Gel 60 F254, Kenilworth, NJ, USA). The compounds were visualized by exposure to UV light
or by dipping the plates into solutions of KMnO4 or vainilline stains followed by heating. HPLC
analyses were performed on a Waters 2695 Instrument (Milford, MA, USA), equipped with a Waters
996 Photodiode Array Detector (Milford, MA, USA). To determine the enantiomeric excesses of diols
(R)-
1
–
6d and acetates (S)-
1
–
6e, the following columns from Daicel (Tokyo, Japan) were employed:
0.46 cm). The optical purity of the diols
Chiralcel OD (25
×
0.46 cm) and Chiralpak AD-H (25
×
was measured after their derivatization to the corresponding acetates using acetic anhydride and
pyridine in CH2Cl2. HPLC conditions and retention times are summarized in Table S1. The absolute
configuration of the 1,2-diols (R)-1–6d and the acetates (S)-1–6e were established by comparison with
the described values of the specific rotation for (R)-ethyl 2-benzyl-2,3-dihydroxypropanoate [23].
3.1. General Procedure for the Synthesis of Racemic Azocompounds (±)-1–6b
The corresponding
(8.0 mL) and N-tert-butyl formaldehyde hydrazone (10 mmol) was added. For the ethyl benzoylformate,
catalyst (0.5 mmol, 10 mol%) was added prior to the hydrazone. Reactions were stirred at room
α-ketoester 1–6a (5.0 mmol) was dissolved at room temperature in toluene
I
temperature for 24 h until consumption of the starting material (TLC). The solvent was eliminated
under reduced pressure and the obtained crudes were purified by column chromatography using
toluene/EtOAc mixtures as the eluent in order to obtain the corresponding racemic azocompounds
(±)-1–6b with yields between 80 and 92%.
3.2. General Preparation of the Racemic 1,2-diols (±)-1–6d Starting from Azocompounds (±)-1–6b
The corresponding azocompound (
to 0 C, and HCl 6.0 M (15 mL) was added. The reaction mixture was allowed to warm to room
±
)-
1
–
6b (4.0 mmol) was dissolved in Et2O (35 mL), cooled
◦
temperature and was stirred for 4 h, and then extracted with Et2O (2
The organic layers were dried over Na2SO4 and the solvent was eliminated in vacuo to create the crude
-hydroxyaldehydes ( )- 6c, which were further reduced without purification. Tetrabutylamonium
×
15 mL) and CH2Cl2 (2
×
15 mL).
α
±
1–
borohydride (560 mg, 2.0 mmol) was added to a solution of the obtained aldehyde in CH2Cl2 (15 mL)
and the mixture was stirred at room temperature for 2 h. After evaporation of the organic solvent,
the crude was purified by column chromatography using CH2Cl2/MeOH 97:3 as the eluent, yielding
the racemic 1,2-diols (±)-1-6d (Figure 2).
(
±
)-Ethyl 2,3-dihydroxy-2-phenylpropanoate, (
±
)-1d
:
Yellow pale oil (395 mg, yield 47%).
Spectroscopic data consistent with the literature [29].
1
(
±
)-Methyl 2,3-dihydroxy-2-phenylpropanoate, (
(300 MHz, CDCl3):
J = 10.8 Hz, H3), 4.13 (bs, 1H, OH), 3.77 (s, 3H, H1), 3.68 (d, 1H, J = 10.8 Hz, H30 ), 2.80 (bs, 1H, OH).
13C-NMR (75.4 MHz, CDCl3):
(ppm) 174.2 (C2), 138.0 (C5), 128.5 (C7), 128.2 (C6), 125.3 (C8), 79.7 (C4),
68.3 (C3), 53.5 (C1). HRMS: m/z calcd. for C10H12NaO4 (M + Na+): 219.02626; found: 219.02628.
±
)-2d: Yellow pale oil (384 mg, yield 49%). H-NMR
δ
(ppm) 7.53 (d, 2H, J = 8.0 Hz, H7), 7.33–7.24 (m, 3H, H6 and H8), 4.18 (d, 1H,
δ
(
±
)-Ethyl 2-(4-cyanophenyl)-2,3-dihydroxypropanoate, ( )-3d: Colorless oil (517 mg, yield 55%).
1H-NMR (300 MHz, CDCl3):
(ppm) 7.69 (d, 2H, J = 8.5 Hz, H8), 7.59 (d, 2H, J = 8.5 Hz, H7), 4.29–4.21
(m, 2H, H2), 4.15 (d, 1H, J = 11.4 Hz, H5), 4.07 (bs, 1H, OH), 3.66 (d, 1H, J = 11.4 Hz, H50 ), 3.09 (bs,
1H, OH), 1.25 (t, 3H, J = 7.1 Hz, H1). 13C-NMR (75.4 MHz, CDCl3):
(ppm) 172.6 (C3), 143.2 (C6),
±
δ
δ

Molecules 2018, 23, 1585
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132.2 (C8), 126.6 (C7), 118.5 (C10), 112.3 (C9), 79.4 (C4), 68.2 (C5), 63.3 (C2), 14.0 (C1). HRMS: m/z calcd.
for C12H13NNaO4 (M + Na+): 258.0739; found: 258.0737.
(
±
)-Ethyl 2,3-dihydroxy-2-(4-methoxyphenyl)propanoate, (
±
)-4d: Colorless oil (432 mg, yield 45%).
Spectroscopic data consistent with the literature [29].
◦
(
±
)-Ethyl 2,3-dihydroxy-2-(tiophen-2-yl)propanoate, (
±
)-5d: White solid. m.p.: 76–78 C (354 mg,
yield 41%). Spectroscopic data consistent with the literature [29].
(
±
)-Ethyl 2-hydroxy-2-hydroxymethyl-4-phenylbutanoate, ( )-6d: Colorless oil (384 mg, yield 57%)
±
1H-NMR (300 MHz, CDCl3):
δ
(ppm) 7.20–7.12 (m, 2H, H10), 7.10–7.07 (m, 3H, H9 and H11), 4.15 (q,
2H, J = 7.0 Hz, H2), 3.98 (s, OH), 3.73 (d, 1H, J = 11.2 Hz, H5), 3.56 (d, 1H, J = 11.2 Hz, H50 ), 2.91 (s,
OH), 2.78–2.68 (m, 1H, H6), 2.43–2.33 (m, 1H, H60 ), 1.98–1.78 (m, 2H, H7), 1.22 (t, 3H, J = 7.0 Hz, H1).
13C-NMR (75.4 MHz, CDCl3):
δ (ppm) 175.0 (C3), 141.2 (C8), 128.5 (C10), 128.4 (C9), 126.0 (C11), 78.3
(C4), 68.0 (C5), 62.4 (C2), 36.6 (C6), 29.5 (C7), 14.2 (C1). HRMS: m/z calcd. for C13H18NaO4 (M + Na+):
261.1102; found: 261.1097.
Figure 2. Structure and NMR assignation of the synthesized racemic 1,2-diols (±)-2,3d and 6d.
3.3. General Synthesis of the Racemic Acetates (±)-1–6e
To a solution of the corresponding racemic 1,2-diol (
pyridine (18 L, 0.22 mmol) and acetic anhydride (20 L, 0.22 mmol) were added at room temperature.
The reaction was stirred until disappearance of the starting material (TLC using hexane/EtOAc 7:3 as
the eluent). The crude reaction was washed with HCl 1.0 N (2 2.0 mL), dried with Na2SO4, and the
solvent was removed under reduced pressure to yield the corresponding racemic acetates ( )- 6e
±
)-1-6d (0.2 mmol) in CH2Cl2 (2.0 mL),
µ
µ
×
±
1– ,
which were obtained after purification by column chromatography using n-hexane/EtOAc 7:3 as
eluent (Figure 3).
(
±
)-Ethyl 3-acetoxy-2-hydroxy-2-phenylpropanoate, (
1H-NMR (300 MHz, CDCl3):
(ppm) 7.57–7.54 (m, 2H, H10), 7.31–7.27 (m, 3H, H9 and H11), 4.68–4.64
(d, 1H, J = 11.3 Hz, H5), 4.30–4.15 (m, 3H, H2 and H50 ), 3.84 (bs, 1H, OH), 2.00 (s, 3H, H7), 1.22 (t,
3H, J = 7.1 Hz, H1); 13C-NMR (75.4 MHz, CDCl3):
(ppm) 172.8 (C6) 170.6 (C3), 137.6 (C8), 129.3
±
)-1e: Colorless oil (45.8 mg, yield 91%).
δ
δ
(C10), 128.5 (C9), 125.6 (C11), 79.2 (C4), 68.8 (C2), 62.9 (C5), 20.7 (C7), 14.1 (C1). HRMS: m/z calcd. for
C13H16NaO5 (M + Na+): 275.0892; found: 275.0890.
(
±
)-Methyl 3-acetoxy-2-hydroxy-2-phenylpropanoate, (
1H-NMR (300 MHz, CDCl3):
(ppm) (ppm) 7.55 (d, 2H, J = 8.2 Hz, H9), 7.31–7.27 (m, 3H, H8 and
H10), 4.64 (d, 1H, J = 11.2 Hz, H4), 4.31 (d, 1H, J = 11.2 Hz, H40 ), 3.77 (s, 3H, H1), 2.54 (bs, 1H, OH), 2.01
±
)-2e: Colorless oil (44.2 mg, yield 93%).
δ
δ
(s, 3H, H6). 13C-NMR (75.4 MHz, CDCl3):
(C8), 125.6 (C10), 77.6 (C3), 68.8 (C4), 53.5 (C1), 20.8 (C6). HRMS: m/z calcd. for C12H14NaO5 (M + Na+):
δ (ppm) 173.2 (C5), 170.6 (C2), 137.5 (C7), 128.7 (C9), 128.5
261.0373; found: 261.0377.
(
±
)-Ethyl 3-acetoxy-2-(4-cyanophenyl)-2-hydroxypropanoate, (
88%). 1H-NMR (300 MHz, CDCl3):
(ppm) 7.74–7.71 (d, 2H, J = 8.6 Hz, H10), 7.62–7.59 (d, 2H,
J = 8.6 Hz, H9), 4.61 (d, 1H, J = 11.3 Hz, H5), 4.31–4.17 (m, 3H, H2 and H50 ), 4.01 (bs, 1H, OH), 2.00
(s, 3H, H7), 1.23 (t, 3H, J = 7.1 Hz, H1). 13C-NMR (75.4 MHz, CDCl3):
(ppm) 171.7 (C6), 170.4 (C3),
±
)-3e: Colorless oil (48.7 mg, yield
δ
δ

Molecules 2018, 23, 1585
8 of 10
141.0 (C8), 132.2 (C10), 126.8 (C9), 119.0 (C12), 112.5 (C11), 77.3 (C4), 68.5 (C5), 63.5 (C2), 20.7 (C7), 14.0
(C1).HRMS: m/z calcd. for C14H15NNaO5 (M + Na+): 300.0842; found: 300.0849.
(
±
)-Ethyl 3-acetoxy-2-hydroxy-2-(4-methoxyphenyl)propanoate, (
±
)-4e: Colorless oil (50.8 mg, yield
1
90%). H-NMR (300 MHz, CDCl3):
δ (ppm) 7.47 (d, 2H, J = 8.1 Hz, H9), 6.91 (d, 2H, J = 8.1 Hz, H10),
4.58 (d, 1H, J = 11.0 Hz, H5), 4.30–4.15 (m, 3H, H2 and H50 ), 3.97 (bs, 1H, OH), 2.03 (s, 3H, H7), 1.30 (t,
3H, J = 7.0 Hz, H1). 13C-NMR (75.4 MHz, CDCl3):
δ
(ppm) 171.8 (C6), 169.7 (C3), 158.0 (C11), 131.1 (C8),
127.2 (C9), 111.0 (C10), 79.3 (C4), 68.0 (C5), 63.0 (C2), 57.2 (C12), 21.2 (C7), 14.0 (C1). HRMS: m/z calcd.
for C14H18O6 (M+) 282.1103; found: 282.1098.
(
±
)-Ethyl 3-acetoxy-2-hydroxy-2-(tiophen-2-yl)propanoate, (
±
)-5e: Yellow pale oil (41.4 mg, yield
1
81%). H-NMR (300 MHz, CDCl3):
δ (ppm) 7.25 (dd, 1H, J = 5.1, 1.3 Hz, H11), 7.10 (dd, 1H, J = 3.7,
1.3 Hz, H10), 6.98 (dd, 1H, J = 5.1, 3.7 Hz, H9), 4.56 (d, 1H, J = 11.1 Hz, H5), 4.33–4.19 (m, 3H, H2 and
H50 ), 2.91 (bs, 1H, OH), 2.00 (s, 3H, H7), 1.25 (t, 3H, J = 7.0 Hz, H1). 13C-NMR (75.4 MHz, CDCl3):
δ
(ppm) 171.8 (C6), 170.3 (C3), 141.7 (C8), 127.2 (C11), 125.9 (C10), 125.0 (C9), 76.4 (C4), 69.2 (C5), 63.2 (C2),
20.7 (C7), 13.9 (C1). HRMS: calcd. for C11H14NaO4S (M + Na+): 281.0456; found: 281.0454.
(
±
)-Ethyl 2-(acetoxymethyl)-2-hydroxy-4-phenylbutanoate, (
1H-NMR (300 MHz, CDCl3):
(ppm) 7.21–7.18 (m, 2H, H12), 7.13–7.07 (m, 3H, H11 and H13), 4.19–4.08
(m 4H, H2, H5 and H50 ), 3.51 (bs, 1H, OH), 2.79–2.70 (m, 1H, H8), 2.45–2.35 (m, 1H, H80 ), 1.99–1.90 (m,
5H, H7 and H9), 1.21 (t, 3H, J = 7.1 Hz, H1). 13C-NMR (75.4 MHz, CDCl3):
(ppm) 173.9 (C6), 170.5
±
)-6e: Colorless oil (47.6 mg, yield 85%).
δ
δ
(C3), 141.0 (C10), 128.4 (C11), 126.1 (C12), 125.7 (C13), 76.0 (C4), 68.9 (C5), 62.4 (C2), 36.8 (C8), 29.3 (C9),
20.7 (C7), 14.2 (C1). HRMS: m/z calcd. for C15H20NaO5 (M+): 303.1204; found: 303.1203.
O
O
O
6
O
6
5
O
O
7
7
6
5,5'
5,5'
4,4'
HO
HO
8
HO
9
9
8
3
8
O
3
2
7
1
O
O
1
10
11
10
11
9
4
4
3
2
2
1
O
O
O
10
9
9
8
NC
10
10
9
12
1e
2e
3e
(±)-
(±)-
(±)-
O
O
O
6
6
O
O
6
O
7
7
7
5,5'
5,5'
HO
8
HO
8
5,5'
HO
11
9
9
9
3
O
3
O
10
1
1
3
8
O
1
10
11
12
13
4
4
4
2
10
2
2
O
12
11
O
9
S
O
O
11
12
10
4e
5e
6e
(±)-
(±)-
(±)-
Figure 3. Structure and NMR assignation of the synthesized racemic acetates (±)-1–6e.
3.4. General Procedure for the Biocatalyzed Acylation of the Racemic 1,2-diols (±)-1–6d
Unless otherwise stated, vinyl acetate (0.9 mmol) was added to a solution of the racemic diol
(
±
)-
Reactions were stirred at 30 ◦C at 220 rpm and monitored by TLC using n-hexane/EtOAc 7:3 as the
eluent. Once finished, the lipase was filtered, washed with TBME (2 2 mL), and the solvent was
evaporated under reduced pressure. The crude mixture was purified by column chromatography
using n-hexane/EtOAc 7:3 as the eluent in order to obtain the (R)-1,2-diols 6d and the (S)-acetates
1–6d (0.3 mmol) in TBME (2.0 mL) containing the PSL-C (30 mg) and Na2CO3 (0.25 mmol).
×
1–
1
–
6e, which were analysed by HPLC at the conditions described in Table S1 for the determination
21
D
of the optical purities. (R)-1d: 37.8 mg, 60% yield; [
mg, 32% yield; [
CHCl3, 55% ee); (S)-2e: 24.1 mg, 35% yield; [
α
]
=
−
2.9 (c = 0.6, CHCl3, 47% ee); (S)-1e: 23.0
21
α
]
21 = +10.8 (c = 1.0, CHCl3, 92% ee). (R)-2d: 33.8 mg, 57% yield; [
α
]
=
−
3.8 (c = 0.5,
D
D
21
α
]
= +12.1 (c = 0.8, CHCl3, 92% ee). (R)-3d: 50.5 mg, 71%
D

Molecules 2018, 23, 1585
9 of 10
21
21
D
yield; [
α
]
=
−
4.2 (c = 1.0, CHCl3, 32% ee); (S)-3e: 19.4 mg, 22% yield; [
α
]
= +9.8 (c = 1.2, CHCl3, 93%
D
21
21
ee). (R)-4d: 51.6 mg, 72% yield; [
α
]
=
−
1.7 (c = 0.4, CHCl3, 30% ee); (S)-4e: 16.9 mg, 20% yield; [
α
]
=
D
D
+6.8 (c = 1.2, CHCl3, 92% ee). (R)-5d: 38.1 mg, 59% yield; [α]2D1 = −2.7 (c = 0.75, CHCl3, 50% ee); (S)-5e:
21
D
21
D
23.2 mg, 30% yield; [
α
]
= +10.2 (c = 1.05, CHCl3, 90% ee). (R)-6d: 34.0 mg, 48% yield; [
α
]
=
−
3.8
(c = 0.5, CHCl3, 50% ee); and (S)-6e: 33.6 mg, 40% yield, [α]2D1 = +5.6 (c = 1.2, CHCl3, 62% ee).
4. Conclusions
A set of aromatic and non-aromatic 1,2-diols containing an ester moiety were prepared in a
three-step procedure with moderate yields from the corresponding -ketoesters. These functionalized
α
racemic 1,2-diols were tested in lipase-catalysed acetylations. After optimization of the reaction
conditions, we achieved good activities and selectivities in the resolution of aromatic 1,2-diols
by employing the Pseudomonas cepacia lipase (PSL-C) in tert-butyl methyl ether as the solvent.
This biocatalyst showed a higher selectivity for the preparation of chiral (R)-1,2-diols and (S)-acetates
containing unsubstituted aromatic rings or presenting electron-withdrawing groups, whereas the
reactions were slower and slightly less selective for aromatic substrates with electron-donating groups
or heteroaromatic systems. PSL-C could be recycled for five reactions without appreciable loss in its
biocatalytic properties, thus resulting in a promising biocatalyst for the preparation of optically active
1,2-diols.
Supplementary Materials: The following are available online at http://www.mdpi.com/1420-3049/23/7/1585/
s1, Table S1: HPLC analyses; and assignment of 1H and 13C-NMR of synthesized compounds.
Funding: This research received no external funding.
Acknowledgments: This work was supported by the Ministerio de Economía y Competitividad, MINECO
(contract RYC-2012-10014 for G.d.G., Grants CTQ2016-76908-C2-1-P and CTQ2016-76908-C2-2-P), the European
FEDER Funds, and the Junta de Andalucía (Grant 2012/FQM 10787).
Conflicts of Interest: Author declares no conflict of interest.
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Sample Availability: Samples of the compounds (±)-1–6d and (±)-1–6e are available from the authors.
©
2018 by the author. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
(CC BY) license (http://creativecommons.org/licenses/by/4.0/).