Synthesis of pyrazine via chemoselective reduction of β-keto-α-oximino ester using baker's yeast

Article abstract of DOI:10.1016/j.molcatb.2015.11.003

The synthesis of pyrazines by the baker's yeast-mediated reaction of β-keto-α-oximino esters and amides is described. Baker's yeast reduced oximes selectively over ketones of β-keto-α-oximino esters to give the corresponding β-keto-α-amino ester intermediates, which underwent spontaneous dimerization followed by air-induced aromatization to yield pyrazines. The chemoselective reduction of β-keto-α-oximino amides using baker's yeast also afforded the corresponding pyrazines. Interestingly, both hydroximes and alkoximes gave the pyrazines by the baker's yeast-mediated reduction via the corresponding amino ketones, the known precursors of pyrazines. The reaction was strongly dependent upon pH of reaction medium, and gave optimum yields at pH 5. These results demonstrate that pyrazines were synthesized efficiently and eco-friendly using a whole-cell biocatalytic system as an alternative to chemical reduction.

Full text of DOI:10.1016/j.molcatb.2015.11.003

Accepted Manuscript
Title: Synthesis of Pyrazine via Chemoselective Reduction of
␤-Keto-␣-Oximino Ester using Baker’s Yeast
Author: Kilwoong Mo Jin Hyeong Park Soon Bang Kang
Youseung Kim Yong Sup Lee Jae Wook Lee Gyochang Keum
PII:
S1381-1177(15)30096-5
DOI:
Reference:
http://dx.doi.org/doi:10.1016/j.molcatb.2015.11.003
MOLCAB 3267
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Journal of Molecular Catalysis B: Enzymatic
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24-10-2015
1-11-2015
Please cite this article as: Kilwoong Mo, Jin Hyeong Park, Soon Bang
Kang, Youseung Kim, Yong Sup Lee, Jae Wook Lee, Gyochang Keum,
Synthesis of Pyrazine via Chemoselective Reduction of rmbeta-Keto-rmalpha-
Oximino Ester using Baker’s Yeast, Journal of Molecular Catalysis B: Enzymatic
http://dx.doi.org/10.1016/j.molcatb.2015.11.003
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Synthesis of Pyrazine via Chemoselective Reduction of β-Keto-α-
Oximino Ester using Baker’s Yeast
Kilwoong Mo,a,b Jin Hyeong Park b,d, Soon Bang Kang,b Youseung Kim,b Yong Sup Lee,a Jae
Wook Lee,*c,d Gyochang Keum*b,d
a Department of Pharmacy, College of Pharmacy & Department of Life and
Nanopharmaceutical Sciences, Kyung Hee University, 1 Hoegi-dong, Dongdaemun- gu,
Seoul 130-701, Republic of Korea
b Center for Neuro-medicine, Korea Institute of Science and Technology (KIST), Hwarang-
ro 14-gil 5, Seongbuk-gu, Seoul, 136-791, Republic of Korea
c Natural Product Research Center, KIST, Saimdang-ro 679, Gangneung, Gangwon-do,
210-340, Republic of Korea
d Department of Biological Chemistry, University of Science and Technology (UST), 217
Gajeong-ro Yuseong-gu, Daejeon 305-350, Republic of Korea
* Corresponding author. Tel:+82 33 650 3514; fax: +82 33 650 3529
E-mail address: jwlee5@kist.re.kr
* Corresponding author. Tel:+82 2 958 5148; fax: +82 2 958 3529
E-mail address: gkeum@kist.re.kr
1

Graphical abstract:
2

Highlights:
 Pyrazines were prepared from β-keto-α-oximino esters using baker’s yeast.
 Baker’s yeast reduced oxime selectively over ketone.
 β-Keto-α-amino ester might be a key intermediate toward pyrazines.
 β-Keto-α-oximino amide reacted with baker’s yeast to give pyrazines too.
 Pyrazines were prepared efficiently and eco-friendly by biocatalyzed reaction
3

Abstract: The synthesis of pyrazines by the baker’s yeast-mediated reaction of β-keto-α-
oximino esters and amides is described. Baker’s yeast reduced oximes selectively over
ketones of β-keto-α-oximino esters to give the corresponding β-keto-α-amino ester
intermediates, which underwent spontaneous dimerization followed by air-induced
aromatization to yield pyrazines. The chemoselective reduction of β-keto-α-oximino amides
using baker’s yeast also afforded the corresponding pyrazines. Interestingly, both hydroximes
and alkoximes gave the pyrazines by the baker’s yeast-mediated reduction via the
corresponding amino ketones, the known precursors of pyrazines. The reaction was strongly
dependent upon pH of reaction medium, and gave optimum yields at pH 5. These results
demonstrate that pyrazines were synthesized efficiently and eco-friendly using a whole-cell
biocatalytic system as an alternative to chemical reduction.
Keywords: Baker’s yeast • Reduction • Chemoselectivity • Pyrazine • β-Keto-α-oximino
esters
4

1. Introduction.
Baker’s yeast (Saccharomyces cerevisiae) is a convenient whole-cell biocatalytic system
that has been used extensively for the asymmetric reduction of a variety of ketone and
carbonyl functionalities [1-5]. Many researchers have shown that baker’s yeast is an efficient
bio-catalyst for the reduction of C-C double bonds,[6] hydrolysis,[7] oxidation,[8] C-C bond
formation,[9] and nitro group reduction.[10] In addition, baker’s yeast has been shown to
reduce oximes to amines [11]. Hydroxylamines have also been formed from oximes through
the reduction of C-N double bonds using an immobilized baker’s yeast in a hexane-water
(1:9) solvent system [12]. The baker’s yeast-mediated reduction of oximes into alcohols or
amines depends on the relative rates of reaction between the hydrolysis of an oxime to its
corresponding carbonyl group and the reduction of the oxime to an amine [13]. Interestingly,
Kreutz et al.[14,15] reported that α-oximino ketones, which have both oxime and ketone
groups, can be reduced into optically active α-hydroxyoximes by baker's yeast (Figure 1).
This stereoselective reduction of ketone by baker’s yeast is influenced by the size of the R
group on the various substrates.
In the chemical reduction, the selectivity towards oxime and carbonyl groups can be
controlled by selecting the appropriate reducing agent. The ketones in β-keto-α-alkyloximino
esters can be selectively reduced to alcohols by sodium borohydride (NaBH4) [16]. The
oxime groups in β-keto-α-hydroximino esters can be selectively reduced to amines by
aluminum amalgam, Zn, Pd/C, SnCl2, or indium [17–23]. The selective reduction of oximes
in β-keto–α-hydroximino esters is useful for stereoselective syntheses of threonine analogues
and natural products [19, 24–27]. Furthermore, pyrazine can be synthesized via the oxime-
selective reduction of α-hydroximino-β-keto esters with aluminum amalgam,[18] zinc,[20]
Raney nickel,[28] Pd/C,[29] or excess aqueous TiCl3,[30] followed by condensation and
aromatization. However, to the best of our knowledge, alkyloxime selective reduction over
ketone of β-keto-α-alkyloximino ester was not reported previously even by chemical reducing
agents.
Pyrazines are nitrogen-containing, six-membered heterocyclic compounds with a long
history in organic chemistry[31] and have been utilized in a broad range of academic and
industrial fields including materials science,[32] food science,[33] and pharmaceutical
research.[34] Pyrazines are found in fermented and heated foods and are widely used in the
food industry as flavor ingredients [33, 35]. They are also found in many vegetables, insects,
and microorganisms [36–39]. Many pyrazine derivatives boast various pharmacological
5

activities including antimicrobial,[40–43] herbicidal,[44] cardiovascular,[45–46] anti-
inflammatory,[47] antidepressant,[48] analgesic,[49–51] and anticancer activities [52–54].
Biosynthesis[55–56] and other synthetic approaches[31,57-59] for pyrazines have been
widely studied because of the broad application base of pyrazine scaffolds. We recently
reported the baker’s yeast-mediated, ketone-selective reduction of β-keto-α-oximino nitriles
to give β-hydroxy-α-oximino nitriles with high stereoselectivity [60]. However, the oxime-
selective reduction of α-oximinoketone provides the corresponding α-aminoketone, which is
a known precursor in the chemical preparation of pyrazine. Baker’s yeast, which has the
ability to reduce both ketones and oximes, can be used to synthesize pyrazines from α-
oximinoketones. These reactions can be also applied to the enantioselective synthesis of β-
hydroxy-α-oximino esters and threonine-class amino acid derivatives, as shown in Figure 1.
Therefore, this study explores the baker’s yeast-mediated selective reduction of β-keto-α-
oximino esters.
2. Materials and methods
2.1. Yeast
Baker's yeast (BY) was purchased from Sigma-Aldrich Co. (St. Louis, MO, USA) and
Oriental (Tokyo, Japan), and stored in a refriderator at 2–8 oC.
2.2. Chemicals
All chemicals and solvents were obtained from commercial suppliers (Fisher Scientific Co.
and Sigma-Aldrich Co.) and used without further purification. Unless otherwise indicated,
biocatalyzed reactions were performed under shaking incubator with temperature control.
2.3. Synthesis of β-keto-α-alkyloximino esters (1b–f) and amide (4b)
Ethyl 2-methoxyimino-3-oxobutanote (1b): To a stirred solution of 1a (2.0 g, 12.6 mmol)
and dimethyl sulfate (1.43 mL, 15.1 mmol) in acetone (20 mL), K2CO3 (1.04 g, 7.56 mmol)
was added slowly at 0 oC. The reaction mixture was stirred at room temperature for 5 h. After
completion of the reaction, monitored by TLC, the reaction mixture was filtered and the
filtrate was concentrated under reduced pressure. The residue was purified by flash
chromatography on silica gel (n-hexane/EtOAc = 20:1) to afford the desired product 1b (2.18
g, 98% yield). 1H NMR (300 MHz, CDCl3) : 4.35 (q, J = 7.14 Hz, 2H), 4.10 (s, 3H), 2.40 (s,
6

3H), 1.33 (t, J = 7.14 Hz, 3H) ppm. 13C NMR (75 MHz, CDCl3) : 193.2, 161.5, 150.4, 64.7,
62.5, 25.5, 14.4 ppm. MS (EI): m/z 173 [M+]. TLC: Rf 0.31 (n-hexane/EtOAc 20:1).
Ethyl 2-(benzyloxyimino)-3-oxobutanoate (1c): The title compound was obtained from
1a and benzyl bromide according to the similar procedure by the O-alkylation described
above. Yield: 87.4%. 1H NMR (300 MHz, CDCl3) : 7.40–7.25 (m, 5H), 5.31 (s, 2H), 4.34 (q,
J = 7.14 Hz, 2H), 2.36 (s, 1H), 1.30 (q, J = 7.15 Hz, 3H) ppm. 13C NMR (75 MHz, CDCl3) :
193.3, 161.6, 150.9, 136.3, 129.0, 128.9, 128.6, 79.0, 62.5, 25.6, 14.4 ppm. MS (EI): m/z 249
[M+]. TLC: Rf 0.28 (n-hexane/EtOAc 20:1).
Methyl 2-(methoxyimino)-3-oxobutanoate (1d): The title compound was obtained from
methyl 2-(hydroxyimino)-3-oxobutanoate according to the same procedure by the O-
methylation described above. Yield: 92%. 1H NMR (300 MHz, CDCl3) : 4.09 (s, 3H), 3.85
(s, 3H), 2.38 (s, 3H) ppm. 13C NMR (75 MHz, CDCl3) : 192.9, 161.7, 150.1, 64.6, 52.8, 25.3
ppm. TLC: Rf 0.28 (n-hexane/EtOAc 20:1).
Methyl 2-(methoxyimino)-3-oxopentanoate (1e): The title compound was obtained from
methyl 2-(hydroxyimino)-3-oxopentanoate according to the same procedure by the O-
methylation described above. Yield: 94%. 1H NMR (300 MHz, CDCl3) : 4.08 (s, 3H), 3.86
(s, 3H), 2.81 (q, J = 7.8 Hz, 2H), 1.12 (t, J = 7.8, 2H) ppm. 13C NMR (75 MHz, CDCl3) :
195.9, 161.9, 149.5, 64.5, 52.8, 31.1, 7.7 ppm. TLC: Rf 0.26 (n-hexane/EtOAc 20:1).
Benzyl 2-(methoxyimino)-3-oxobutanoate (1f): The title compound was obtained from
benzyl 2-(hydroxyimino)-3-oxobutanoate according to the same procedure by the O-
methylation described above. Yield: 94%. 1H NMR (300 MHz, CDCl3) : 7.40–7.37 (m, 5H),
5.33 (s, 2H), 4.10 (s, 3H), 2.41 (s, 3H) ppm. 13C NMR (75 MHz, CDCl3) : 192.9, 161.3,
150.0, 134.9, 128.8, 128.7, 128.4, 67.7, 64.6, 25.4 ppm. MS (EI): m/z 235 [M+]. TLC: Rf 0.31
(n-hexane/EtOAc 15:1).
2-(Methoxyimino)-3-oxo-N-phenylbutamide (4b): The title compound was obtained from
4a according to the same procedure by the O-methylation described above. Yield: 97%. 1H
NMR (300MHz, CDCl3) : 7.50–6.90 (m, 5H), 4.07 (s, 3H), 2.38 (s, 3H) ppm. 13C NMR (75
MHz, CDCl3) : 194.5, 155.5, 150.9, 137.1, 129.5, 129.4, 125.5, 121.2, 120.6, 65.1, 26.4 ppm.
MS (EI): m/z 220 [M+]. TLC: Rf 0.38 (n-hexane/EtOAc 10:1).
2.4. Synthesis of pyrazines (3, 3a–c and 6) using Baker’s yeast
7

Diethyl 3,6-dimethylpyrazine-2,5-dicarboxylate (3): Ethyl-2-methoxyimino-3-
oxobutanote 1b (69.2 mg, 0.4 mmol) dissolved in ethanol (1.5 mL) was added to a
suspension of baker’s yeast (2.0 g) and saccharose (3.0 g) in tap water (60 mL) with shaking
at 30 oC. The mixture was shaken for the specified time by monitoring the complete
consumption of the starting material with TLC, and then saturated with sodium chloride. The
resulting mixture was filtered through a celite pad and washed with chloroform. The filtrate
was extracted with chloroform three times, and the organic layer was dried over Na2SO4 and
concentrated under reduced pressure. The residue was purified by flash chromatography on
silica gel (n-hexane/EtOAc = 5:1) to afford the desired product 3 (26.4 mg, 53% yield). 1H
NMR (300 MHz, CDCl3) : 4.02 (s, 6H), 3.12 (q, J = 7.47 Hz, 4H), 1.32 (t, J = 7.47 Hz, 6H)
ppm. 13C NMR (75 MHz, CDCl3) : 165.5, 151.2, 144.5, 62.8, 22.7, 14.6 ppm. HRMS (EI):
m/z calcd for C12H16N2O4: 252.1110 [M+]; found: 252.1151; elemental analysis calcd (%) for
C12H16N2O4: C 57.13, H 6.39, N 11.10, O 25.37; found: C 57.95, H 6.46, N 10.58, O 25.01.
TLC: Rf 0.44 (n-hexane/EtOAc 5:1).
Dimethyl 3,6-dimethylpyrazine-2,5-dicarboxylate (3a): The title compound was
obtained from 1d according to the similar procedure using the baker’s yeast described above.
Yield: 65%. 1H NMR (300 MHz, CDCl3) : 4.03 (s, 6H), 2.84 (s, 6H) ppm. 13C NMR (75
MHz, CDCl3) : 165.6, 151.6, 143.9, 53.6, 22.7 ppm. MS (FAB): m/z 225.22 [M+H]+;
elemental analysis calcd (%) for C10H12N2O4: C 53.57, H 5.39, N 12.49, O 28.54; found: C
53.94, H 5.54, N 11.95, O 27.82. TLC: Rf 0.32 (n-hexane/EtOAc 10:1).
Dimethyl 3,6-diethylpyrazine-2,5-dicarboxylate (3b): The title compound was obtained
from 1e according to the similar procedure using the baker’s yeast described above. Yield:
24%. 1H NMR (300 MHz, CDCl3) : 4.01 (s, 6H), 3.11 (q, J = 7.5 Hz, 4H), 1.26 (t, J = 7.4
Hz, 6H) ppm. 13C NMR (75 MHz, CDCl3) : 167.9, 156.2, 144.4, 51.5, 28.9, 14.5 ppm. MS
(FAB): m/z 253.25 [M+H]+. TLC: Rf 0.31 (n-hexane/EtOAc 10:1).
Dibenzyl 3,6-dimethylpyrazine-2,5-dicarboxylate (3c): The title compound was obtained
from 1f according to the similar procedure using the baker’s yeast described above. Yield:
37%. 1H NMR (300 MHz, CDCl3) : 7.48–7.26 (m, 10H), 5.45 (s, 4H), 2.75 (s, 6H) ppm. 13C
NMR (75 MHz, CDCl3) : 165.2, 151.3, 144.3, 135.4, 129.1, 129.0, 128.9, 68.3, 22.7 ppm.
MS (FAB): m/z 377.16 [M+H]+. TLC: Rf 0.43 (n-hexane/EtOAc 10:1).
3,6-dimethyl-N2,N5-diphenylpyrazine-2,5-dicarboxamide (6): The title compound was
obtained from 4a according to the similar procedure using the baker’s yeast described above.
8

Yield: 45%. 1H NMR (300 MHz, CDCl3) : 9.96 (s, 2H), 7.77–7.16 (m, 10H), 3.10 (s, 6H)
ppm. 13C NMR (75 MHz, CDCl3) : 161.7, 151.3, 143.2, 137.8, 129.5, 125.1, 120.3, 23.1
ppm. MS (FAB): m/z 347.24 [M+Na]+. TLC: Rf 0.32 (n-hexane/EtOAc 5:1).
3. Result and discussion
Ethyl-2-(hydroxyimino)-3-oxobutanoate 1a was used as a substrate to evaluate the baker’s
yeast-mediated reduction of β-keto-α-oximino esters. This reaction using Sigma type II yeast
under typical conditions [60] at 30 oC for 3.5 days afforded a white solid in a 42% yield after
isolation. The solid product stained lightly on a TLC plate, exhibited an unexpectedly low
polarity, and was identified as pyrazine 3. Also, GC/MS analysis confirmed the presence of
pyrazine with a small amount of α-amino ketone intermediate in the reaction mixture.
Interestingly, GC/MS analysis of crude product revealed none of the expected side products
such as compound 2. This result indicated that baker’s yeast selectively reduced the oxime in
1a to give the corresponding α-amino ketone intermediate followed by the spontaneous
transformation to the pyrazine derivative [17–23, 28–30].
To improve the yield of pyrazine 3, the reaction was evaluated under a variety of conditions
by changing additives, temperature, and the type of baker’s yeast used (table 1). The
reduction with baker’s yeast immobilized on montmorillorite K10[15] gave a similar yield as
that obtained with entry 1. Reductions without saccharose gave lower yields with a mixture
of unknown compounds (entry 3). Further modification of the reaction conditions did not
improve the isolation yield of compound 3.
In addition to hydroximes, alkyloximes were also evaluated as substrates for baker’s yeast-
mediated, oxime-selective reduction. A series of β-keto-α-alkyloximino esters 1b and 1c were
prepared according to a previous method [25]. Treatment of the appropriate β-ketoesters with
sodium nitrite in acetic acid gave hydroxime compounds 1a and subsequent O-alkylation of
the hydroximes gave the methyl- and benzyl-oximinoketones 1b and 1c, respectively, in high
yields. The reduction of α-methyloximinoketone 1b with a variety of baker’s yeasts gave
pyrazine 3 with relatively low yields. GC/MS analyses showed the presence of small amounts
of alcohol 2 generated by β-ketone reduction. The results represent that alkyloxime groups in
β-keto-α-alkyloximino esters can be selectively reduced to α-aminoketones by baker’s yeast.
The reduction reaction in the absence of saccharose slightly improved the isolated yield of
3 (31%) following relatively long reaction times (3.5 days, entry 9). β-Keto-α-benzyloximino
9

ester 1c with Sigma type II baker’s yeast successfully afforded pyrazine 3 in a 38% isolated
yield with a 9:91 GC ratio of 2:3, which is a higher selectivity than that obtained with the
immobilized yeast (entries 11 and 12). The reaction times of alkyloximes 1b and 1c were
shorter than that of the corresponding hydroxime 1a.
When a variety of reaction conditions were evaluated, the results in table 2 show that the
reaction was primarily influenced by pH. The reaction at pH 7 showed a significantly lower
isolation yield after 1.5 days (entry 1, table 2) than that obtained without buffer. The product
yield increased with decreasing pH until reaching an optimum at pH 5 (entry 3).
These results show that an addition of small amount of ethanol and careful control of the
reaction pH (entries 1–4) range are critical to improving yields in baker’s yeast-mediated
reductions of 1a. The reductions of methyl oxime 1b also gave pyrazine 3 in high yields at
pH 5 with three types of baker’s yeast. The reduction of benzyl oxime 1c showed the highest
yield with Sigma type I baker’s yeast at the same pH (entries 9–11). The reactions of methyl
and benzyl esters 1d–f, prepared using the same method, and the corresponding pentanoate
1e also afforded the chemoselective reduction of the oxime to afford pyrazines 3a–c (entries
12–14 in table 2). The reduction of methyl ester 1d with baker's yeast purchased from
Oriental showed the highest isolation yield (65%, entry 13 in table 2).
To test β-keto-α-oximino amide substrates, we prepared compounds 4a–b from 2,2,6-
trimethyl-4H-1,3-dioxin-4-one and aniline using a known procedure [61]. The reaction of
hydroxime amide 4a gave a 1:2 mixture of diol 5 and pyrazine 6 in GC analyses in 9% and
21% isolated yields, respectively, using Sigma type II baker’s yeast without buffer (entry 1 in
table 3).
The same reaction of 4a at pH 5 gave pyrazine 6 with improved yield (45%, entry 2 in table
3). The reduction of methyloxime amide 4b with baker’s yeast also performed best at pH 5
and yielded pyrazine 6 in a low yield of 25%. With the exception of the benzyl ester 1f,
reactions with N-phenyl amides 4a and 4b required longer durations than reactions with
methyl and ethyl esters 1a–e (table 3).
We next explore the mechanism of formation of compound 3 (dimethyl 3,6-
dimethylpyrazine-2,5-dicarboxylate) by baker’s yeast mediated reduction, and the plausible
mechanism was shown in Figure 2. Usually, the product which is generated by baker’s yeast
-ketone was expected to be alcohol. The GC-MS analysis revealed that the
molecular mass of amino ketone was found in the reaction mixture confirming the generation
of amino ketone 7 as an intermediate. The baker's yeast reduced oximino ketone to amino
10

ketone. The amino ketone 7 were dimerized by spontaneous condensation and followed by air
oxidation to give pyrazine.
4. Conclusion
In summary, the synthesis of pyrazines through the baker’s yeast-mediated reaction of β-
keto-α-oximino esters and amides was developed. The pyrazines were generated by the
oxime-selective reduction of β-keto-α-oximino esters and amides with baker’s yeast to their
corresponding aminoketone intermediates, which was not isolated but underwent spontaneous
dimerization followed by air-oxidized aromatization to provide pyrazines. The selective
reduction with baker’s yeast converted both the hydroxy and alkyl oxime moieties of β-keto-
α-oximino esters and amides to the corresponding β-keto-α-amino esters and amides, the
known precursors of pyrazines. The baker’s yeast-mediated reduction was primarily
influenced by pH and gave the optimum results at pH 5. The results demonstrate that
pyrazines can be prepared in an efficient and eco-friendly green procedure using a whole-cell
biocatalytic system, representing a valuable alternative to chemical reduction. Furthermore,
the reactions studied herein may be representative of the processes occurring in biological
systems and may account for the natural occurrence of pyrazines in foods and
microorganisms.
Acknowledgements
This work was supported by the Korea Institute of Science and Technology (KIST)
institutional program (2E25240 and 2Z04371).
11

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14

Figure 1. Baker’s yeast (BY)-mediated reduction of β-keto-α-oximino carbonyl
compounds.
15

Figure 2. Proposed mechanism of the synthesis of pyrazine by baker's yeast mediated
reduction.
16

Table 1. Reduction of β-keto-α-alkyloximino esters using baker’s yeast.
Conditions
B.Y.b
GC ratio
yield 3
(%)e
42
entry
sub.
time
3.5 d
3.5 d
3.5 d
3.5 d
3.5 d
3.5 d
6 h
smc
0
ukd
0
2
0
3
100
100
51
94
57
79
86
98
96
49
91
84
1
2
Sigma-II
ImS-II
1a
0
0
0
42
f
3
ImS-II
0
0
49
6
-
4
ImS-II
0
0
22g
5
Sigma-I
Oriental
Sigma-II
ImS-II
14
0
0
29
21
-
-
6
0
22
-
7
0
14
2
1b
1c
8
14 h
3.5 d
14 h
18 h
14 h
0
-
29
31f
-
9
ImS-II
0
4
-
10
11
12
Oriental
Sigma-II
ImS-II
0
10
9
41
-
0
38
-
0
16
-
a
Typical conditions: Baker’s yeast (2.0 g), substrate (0.4 mmol), saccharose (3.0 g) in water
b
(30 mL) and ethanol (1.5 mL) at 30 °C. Baker’s yeast: Sigma-II (Sigma type II), Sigma-I
(Sigma type I), ImS-II (Sigma type II immobilized on montmorillorite K10)[15], Oriental
(Tokyo, Japan). c Starting material. d Unknown mixture. e Isolated yields. f Saccharose was not
used. g Reaction temperature: at 37 oC.
17

Table 2. Reduction of various β-keto-α-oximino esters 1a-f with baker’s yeast
under pH control.
Conditions
yield 3
entry sub.
pdt
(%)b
R1
Me
R2
H
R3
Et
B.Y.
time
1.5 d
3.5 d
1.5 d
1.5 d
2 d
1.5 d
1.5 d
2 d
2 d
1.5 d
2 d
3 d
2 d
pH
7
6
5
4
5
5
5
5
5
5
5
5
1
2
1a
Sigma-II
Sigma-II
Sigma-II
Sigma-II
Oriental
Sigma-II
Sigma-I
Oriental
Sigma-II
Sigma-I
Oriental
Oriental
Sigma II
3
3
3
3
3
3
3
3
3
3
3
3a
3b
3
42
47
27
34
53
35
44
29
40
25
65
24
3
4
5
6
7
8
9
1b
1c
Me
Me
Me
Bn
Et
Et
10
11
12
13
1dc
1ec
Me
Et
Me
Me
Me
Me
5
14
1fc
Me
Me
Bn
Oriental
6 d
5
3c
38
a
Typical conditions: Baker’s yeast (2.0 g), substrate (0.4 mmol), and saccharose (3.0 g) in
water (60 mL) and ethanol (1.5 mL) at 30°C. The pH was adjusted with 1 N HCl and NaOH.
b Isolated yields. c The best results from the three types of baker's yeast were reported.
18

Table 3. Reaction of β-keto-α-oximino amide compounds with baker’s yeast.
entry
sub.
Conditions
time (day)
yield (%)
of 6[c]
yeast type
pH
[b]
1
2
3
4
5
Sigma II
Sigma II
Oriental
Sigma II
Oriental
3.5
5
-
21 (9)[d]
45
4a
5
5
5
5
5
37
7
25
4b
3
25
a Reactions were carried out with 0.4 mmol substrate, baker’s yeast (2.0 g), and saccharose (3.0 g) in
water (60 mL) and ethanol (1.5 mL) at 30°C. The pH was adjusted with 1 N HCl and NaOH.
b Reaction was carried out without buffer. c Isolated yields. d The isolated yield of diol 5 is in
parentheses.
19