Effective Hydrogenation of 3-(2”-furyl)- And 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one in Selected Yeast Cultures

Article abstract of DOI:10.3390/molecules24173185

Biotransformations were performed on eight selected yeast strains, all of which were able to selectively hydrogenate the chalcone derivatives 3-(2”-furyl)-(1) and 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (3) into 3-(2”-furyl)- (2) and 3-(2”-thi

Full text of DOI:10.3390/molecules24173185

molecules
Article
Effective Hydrogenation of 3-(2”-furyl)- and
3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one
in Selected Yeast Cultures
Mateusz Łuz˙ny * , Martyna Krzywda, Ewa Kozłowska , Edyta Kostrzewa-Susłow
and Tomasz Janeczko *
Department of Chemistry, Wrocław University of Environmental and Life Sciences, Norwida 25, 50-375 Wrocław, Poland
* Correspondence: mat.luzny@gmail.com(M.Ł.); janeczko13@interia.pl (T.J.); Tel.: +48-713-205-195 (T.J.)
ꢀꢁꢂꢀꢃꢄꢅꢆꢇ
ꢀꢁꢂꢃꢄꢅꢆ
Received: 21 August 2019; Accepted: 31 August 2019; Published: 2 September 2019
Abstract:
which were able to selectively hydrogenate the chalcone derivatives 3-(2”-furyl)-
) and 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one ( ) into 3-(2”-furyl)- ( ) and
3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propan-1-one ( ) respectively. The highest efficiency of
hydrogenation of the double bond in the substrate was observed in the cultures of
Biotransformations were performed on eight selected yeast strains, all of
(
1
3
2
4
1
Saccharomyces cerevisiae KCh 464 and Yarrowia lipolytica KCh 71 strains. The substrate was converted
into the product with > 99% conversion just in six hours after biotransformation started. The compound
containing the sulfur atom in its structure was most effectively transformed by the Yarrowia lipolytica
KCh 71 culture strain (conversion > 99%, obtained after three hours of substrate incubation).
Also, we observed that, different strains of tested yeasts are able to carry out the bioreduction of
the used substrate with different yields, depending on the presence of induced and constitutive ene
reductases in their cells. The biggest advantage of this process is the efficient production of one
product, practically without the formation of side products.
Keywords: biotransformation; chalcone; dihydrochalcone; yeast; sweeteners
1. Introduction
In recent years, there has been growing interest from the food industry in sweeteners. Within this
group of substances, dihydrochalcones are increasingly gaining attention. The increase in interest
is due to the fact that dihydrochalcones are synthesized by plant cells, and they are a daily part
of our diet [
plants [ ]. Dihydrochalcones have a wide spectrum of activity, such as antiviral (the ability to inhibit
dengue virus proteases or herpes simplex virus), and anti-inflammatory and antioxidant activity [ ].
They are also known for their activity against pathogenic microorganisms, including gram-positive
and gram-negative bacteria, as well as fungi [ ] and antimalarial or anti-tuberculosis activity [ ].
Dihydrochalcone (phlorizin) is an active inhibitor of fungal tyrosinase [ 10]. Dihydrochalcones are
1,2]. They are present in citrus, apples, tomatoes, potatoes, bean sprouts, and other
3
4–7
8
9
6,
also used in chemical synthesis to obtain biologically active compounds. 20-Hydroxydihydrochalcon
is used as a building block in the synthesis of propafenone—the active substance of anti-arrhythmic
drugs [11–13].
Schallenberger’s hypothesis explains the sensation of sweet taste, according to which the flavor
substance and the receptor create contact by hydrogen bonding [14]. This is due to the proton donor
groups (AH) and an electron donor (B), which are also present in taste receptors and the structure of
sweet substance. The sweetness of dihydrochalcones is related to their structure. The most important
element of the structure of these compounds is the substitution of the B-ring in the meta or para position
Molecules 2019, 24, 3185; doi:10.3390/molecules24173185
www.mdpi.com/journal/molecules

Molecules 2019, 24, 3185
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with at least one -OH group. The compounds that have three adjacent substituents on the B-ring do
not have a sweet taste [15]. It is suspected that the structure of the B-ring also affects the perception of
non-sweet aftertaste in dihydrochalcones, which do not always correspond to human preferences [16].
Commonly used in the food industry, neohesperidin dihydrochalcone is characterized by a licorice
flavor and a cool feeling on the tongue [17]. There is also a group of dihydrochalcone analogs,
that have a heteroatom in the B-ring and having non-sweet flavors [2]. This group also describes
3-(2”-furyl)-1-(2’-hydroxyphenyl)-propan-1-one (2), which, depending on the concentration, may have
different taste sensations: A concentration of 0.01 ppm does not show any flavor properties; 1 ppm,
soapy; umami, bitter; 10 ppm, licorice, soapy, slightly sweet, bitter, vegetable, terpenic; a particularly
interesting impression is noted in higher concentrations – 100 ppm, bitter, licorice, light-sweet, celery,
umami, and broth [2].
A method for obtaining 3-(2”-furyl)-1-(2’-hydroxyphenyl)-propan-1-one (2) with a conversion
of 75%, as a result of the chemical synthesis of 2’-hydroxyacetophenone and furfuryl alcohol with
NaOH, in the presence of an iridium catalyst, has been identified in [18]. The direct chemical
hydrogenation of chalcones to dihydrochalcones requires the use of metal salts (complexes) of
iridium, palladium, ruthenium, or nickel, and the entire reaction (due to its high flammability)
must be conducted under strictly controlled conditions [19–22]. There are no reports in the
literature regarding the use of biotechnological methods to obtain both, 3-(2”-furyl)- (2) as well
3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propan-1-one (4).
The majority of living organisms are able to hydrogenate the double bond [23
their specific enzymes, capable of hydrogenating chalcones [ 25 26]. Such activity is also described in
relation to other microorganisms, including bacteria, such as Gordonia sp. and Rhodococcus sp. [27],
the entomopathogenic fungus, Aspergillus flavus [ ], and the cyanobacterium, Spirulina platensis [28].
The reduction of activated alkenes by ente reductases [EC 1.3.1.31], which are flavoproteins from the
old yellow enzyme (OYE) family, has been investigated in great detail [29 31]. The substrates in these
,24]. Yeasts possess
9,
,
6
–
reactions are predominantly hydroxy and methoxy derivatives of chalcones. In this study, we aimed
to assess whether the heteroatom-containing substrate in the B-ring would also be accepted by the
double-binding dehydrogenase, present in the tested biocatalysts.
The main aim of the study was to assess the capability of yeast strains for the biotransformation
of a compound, containing a furan (
experiment, are the subject of many years of research conducted in the Department of Chemistry of
Wrocław University of Environmental and Life Sciences, regarding their catalytic capabilities [26 32 33],
and have been selected as having a high ability in reduceing the double bond in the chalcone
and its derivatives [26 28]. An additional goal was to optimize the biotechnological production of
dihydrochalcone ( ), which would enable the development of the method of obtaining the product
on an increased scale.
1) and thiophene (3) substituent. Yeast strains, used in this
,
,
–
2, 4
2. Results and Discussion
Each of the eight tested microorganisms (Rhodotorula rubra KCh 4, Yarrowia lipolytica KCh 71,
Rhodotorula marina KCh 77, Rhodotorula rubra KCh 82, Candida viswanathii KCh 120, Rhodotorula glutinis
KCh 242, Saccharomyces cerevisiae KCh 464, Candida parapsilosis KCh 909) was able to transform both
substrates into the expected products, but the efficiency of the described process differs significantly
between the strains (Table 1 and Figure 1). High regioselectivity of the biocatalysts’ ability to reduce
the double bond was observed, which was described before on the example of other compounds, e.g.,
chalcone containing no substituents and 20-hydroxychalcone [32,34,35].

Molecules 2019, 24, 3185
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Table 1. Relationship between transformation of substrates
1 and 3 [%] and time in the cultures of
tested strains.
Strain Number
KCh 82 KCh 120
Time [Days]
KCh 4
KCh 71
KCh 77
KCh 242
KCh 464
KCh 909
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
7
91
99
>99
23
33
49
98
>99
92
38
60
63
98
>99
>99
59
84
97
49
>99
>99
84
99
99
0
0
17
52
67
87
98
>99
>99
98
99
98
2
14
26
9
12
16
>99 >99 >99
>99 >99 >99
Figure 1. Time dependence of the transformation of chalcone (
KCh 4; (B) Yarrowia lipolytica KCh 71.
1) in the culture of: (A) Rhodotorula rubra
In the experiment, 3-(2”-furyl)- (
) were obtained by chemical synthesis, and then converted to 3-(2”-furyl)- (
3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propan-1-one using biotransformation methods on
a semi-preparative scale (Figures 2 and 3). The obtained products were purified (isolated
1
) and 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one
(3
2
) and
(4)
1–4
yield > 70%) and their structures were determined on the basis of NMR analysis (1H-NMR, 13C-NMR
and correlation spectra – HMBC; HMQC, COSY) as well as by gas chromatography (GC) and thin-layer
chromatography (TLC) analysis (Supplementary Materials).
O
O
1"
6'
A
3'
2"
1'
2'
O
B
O
5'
1
3
microorganism
2
5"
4'
3"
4"
OH
OH
2
1
Figure 2. Biotransformation of 3-(2”-furyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (
1) by selected yeast
cultures. The ring designations and the numbering of carbon atoms have been placed on compound
1.
O
O
S
S
microorganism
OH
OH
4
3
Figure 3. Biotransformation of 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (3) by selected
yeast cultures.

Molecules 2019, 24, 3185
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The reduction of the double bond was the most efficient in the Saccharomyces cerevisiae KCh 464
strain, where over 99% of the product was observed in the reaction medium after six-hour substrate
incubation (Table 2). On the first day of the reaction, the four tested microorganisms exceeded the
threshold of 90% of the substrate conversion: R. rubra KCh 4, R. rubra KCh 82, Y. lipolytica KCh 71,
2
1
R. marina KCh 77. In the culture of C. viswanathii KCh 120, the 90% limit was exceeded after three
days of the biotransformation process. A surprisingly low conversion was observed for the strains
R. glutinis KCh 242 and C. parapsilosis KCh 909 (Table 1). High hydrogenation of the double bond in the
cultures of these strains was observed for chalcone and its methoxy derivatives [32,36].
Table 2. Transformation of the substrate 1 by selected strains.
Strain Number
Time [h]
KCh4
5 ± 1.1
6 ± 0.1
9 ± 0.5
10 ± 0.9
KCh71
52 ± 4.5
88 ± 1.6
>99 ±0.0
>99 ± 0.0
KCh120
2 ± 0.5
2 ± 0.4
4 ± 0.9
9 ± 0.5
KCh464
40 ± 2.1
84 ± 2.9
98 ± 0.5
99 ± 0.6
1
3
6
12
Substrate 3 was most effectively transformed by the Y. lipolytica KCh 71 strain. The conversion
above 99% was recorded after the third hour of biotransformation. A high efficiency of reduction
of the double bond of chalcone derivatives was also confirmed for Saccharomyces cerevisiae KCh 464
strain (conversion > 98% after three hours of transformation). For most of the tested strains, a lower
conversion of the substrate, containing thiophene (
), was observed. Interestingly, the strain Rhodotorula rubra KCh 4 after twelve hours showed only
10% of substrate conversion, and after 24h it was already 91% (Figure 1), which may indicate that ene
3) in its, compared to the furan-containing substrate
(1
1
reductases are not present in the cell, but their production begins as a response to the stimulus (substrate
induction) from the environment. An analogous induction of dehydrogenases (ene reductases) was
observed for the strain C. viswanathii KCh 120. In the culture of this strain, substrate 3 was converted
faster. However, conversion above 98% was achieved for both substrates at the same time in three
days. Based on the obtained results, it can be concluded that the constitutive enzyme is present
in Y. lipolytica KCh 71 and S. cerevisiae KCh 464 strains (observations for both substrates
Biotransformations with these microorganisms show the fastest conversion progress, compared to
other used microorganisms for both substrate and (Tables 1–3). Similar observations were obtained
1 and 3).
1
3
during the biotransformation of chalcone and described earlier [32].
Table 3. Transformation of the substrate 3 by selected strains.
Strain Number
Time [h]
KCh71
KCh120
KCh242
1 ± 0.6
5 ± 0.5
9 ± 0.6
12 ± 0.9
KCh464
35 ± 3.6
64 ± 0.9
83 ± 0.6
98 ± 0.3
1
3
67 ± 3.9
0 ± 0.0
>99 ± 0.0
>99 ± 0.0
>99 ± 0.0
10 ± 1.1
14 ± 1.2
25 ± 3.1
6
12
The strains of the species Saccharomyces cerevisiae are widely described as biocatalysts,
effectively reducing the double bond in various compounds – derivatives of chalcones [ ], containing
methyl, methoxy, hydroxy substituents [37], and even bromine or chlorine [33 38], in both A and
9
,
B-rings. On the basis of the results obtained in this study, it can be concluded that a chalcone derivative,
containing a heteroatom in the B-ring, is effectively transformed by this strain. In previous studies

Molecules 2019, 24, 3185
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during biotransformation of chalcones, in addition to the hydrogenation product, a carbonyl reduction
product was also observed [28 32 37 39]. In this study, additional products were not observed in any of
,
,
,
the reactions carried out for the test compounds.
Biotransformation
The obtained product 3-(2”-furyl)-1-(2’-hydroxyphenyl)-propan-1-one (
2
) was characterized by
1
the following NMR spectrum: H-NMR (600 MHz) (CDCl3)
δ (ppm): 3.07-3.12 (m, 2H, H-3), 3.34-3.41
(m, 2H, H-2), 6.06 (dq, 1H, J = 3.2, 0.8 Hz, H-3”), 6.29 (dd, 1H, J = 3.1, 1.9 Hz, H-4”), 6.90 (ddd, 1H,
J = 8.2, 7.1, 1.2 Hz, H-5’), 7.02 (dd, 1H, J = 8.4, 0.9 Hz, H-3’), 7.32 (dd, 1H, J = 1.8, 0.8 Hz, H-5”),
7.49 (ddd, 1H, J = 8.5, 7.2, 1.6 Hz, H-4’), 7.92 (dd, 1H, J = 8.0, 1.6 Hz, H-6’), 12.23 (s, 1H, -OH).
13C-NMR (151 MHz, CDCl3)
δ = 22.50 (C-3), 36.73 (C-2), 105.69 (C-3”), 110.44 (C-4”), 118.72 (C-3’),
119.12 (C-5’), 119.40 (C-1’), 129.94 (C-6’), 136.56 (C-4’), 141.41 (C-5”), 154.33 (C-2”), 162.56 (C-2’),
204.90 (C-1).
The obtained product 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propane-1-one (
4
) was characterized by
1
the following NMR spectrum: H-NMR (600 MHz) (CDCl3)
δ (ppm): 3.27-3.33 (m, 2H, H-3), 3.37-3.43
(m, 2H, H-2), 6.87 (dq, 1H, J = 3.4, 1.0 Hz, H-3”), 6.90 (ddd, 1H, J = 7.4, 7.1, 1.0 Hz, H-5’), 6.29 (dd,
1H, J = 5.1, 3.4 Hz, H-4”), 6.99 (ddd, 1H, J = 8.4, 1.1, 0.4 Hz, H-3’), 7.14 (dd, 1H, J = 5.1, 1.2 Hz, H-5”),
7.47 (ddd, 1H, J = 8.4, 7.3, 1.5 Hz, H-4’), 7.76 (dd, 1H, J = 8.1, 1.6 Hz, H-6’), 12.24 (s, 1H, -OH).
13C-NMR (151 MHz, CDCl3)
δ = 24.16 (C-3), 40.26 (C-2), 118.73 (C-3’), 119.13 (C-5’), 119.38 (C-1’),
123.72 (C-5”), 124.99 (C-3”), 127.06 (C-4”), 129.90 (C-6’), 136.60 (C-4’), 143.40 (C-2”), 162.56 (C-2’),
204.76 (C-1).
3. Materials and Methods
3.1. Substrate
The substrates, used for biotransformation, were obtained by Claisen-Shmidt condensation
reaction of 2-hydroxyacetophenone
from Sigma-Aldrich (St. Louis, MO, USA)] dissolved in methanol in an alkaline
environment at high temperature (Figures 4 and 5) according to the procedure described
previously 32 40]. The resulting 3-(2”-furyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (1)
(
5
)
with furfural
(6
)
or aldehyde
7
[purchased
[
,
and 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (3) were used as a substrates for
the biotransformation.
O
O
O
O
NaOH, MeOH
O
H
reflux through 2h
+
OH
OH
1
5
6
Figure 4. Synthesis of 3-(2”-furyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (1).
O
O
O
S
NaOH, MeOH
S
H
reflux through 2h
+
OH
OH
3
5
7
Figure 5. Synthesis of 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (3).
The resulting compound (1) was characterized by the following NMR spectral data:

Molecules 2019, 24, 3185
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1H-NMR (600 MHz) (CDCl3)
δ
(ppm): 6.54 (dd, 1H, J = 3.4, 1.8 Hz, H-3”), 6.77 (d, 1H, J = 3.4 Hz,
H-4”), 6.93 (ddd, 1H, J = 8.2, 7.2, 1.1 Hz, H-5’), 7.02 (dd, 1H, J = 8.4, 0.9 Hz, H-3’), 7.49 (ddd, 1H, J = 8.6,
7.2, 1.6 Hz, H-4’), 7.55 (d, 1H, J = 15.1 Hz, H-2), 7.56 (d, 1H, J = 1.4 Hz, H-5”), 7.68 (d, 1H, J = 15.2 Hz,
H-3), 7.92 (dd, 1H, J = 8.1, 1.6 Hz, H-6’), 12.89 (s, 1H, -OH).
13C-NMR (151 MHz, CDCl3)
δ = 113.03 (C-4”), 117.29 (C-3”), 117.73 (C-2), 118.66 (C-3’), 118.98
(C-5’), 120.17 (C-1’), 129.77 (C-6’), 131.26 (C-3), 136.44 (C-4’), 145.55 (C-5”), 151.65 (C-2”), 163.66 (C-2’),
193.46 (C-1).
The resulting compound (3) was characterized by the following NMR spectral data:
1H-NMR (600 MHz) (CDCl3)
δ (ppm): 6.95 (ddd, 1H, J = 8.1, 7.1, 1.1 Hz, H-5’), 7.02 (dd, 1H, J = 8.4,
1.2 Hz, H-3’), 7.12 (ddd, 1H, J = J = 5.0, 3.7, 0,3 Hz, H-4”), 7.41 (d, 1H, J = 3.6, Hz, H-3”), 7.44 (d, 1H,
J = 15.2 Hz, H-2), 7.47 (d, 1H, J = 4.9 Hz, H-5”), 7.49 (ddd, 1H, J = 8.4, 7.2, 1.5 Hz, H-4’), 7.89 (dd, 1H,
J = 8.1, 1.6 Hz, H-6’), 8.05 (dd, 1H, J = 15.2, 0,5 Hz, H-3), 12.85 (s, 1H, -OH).
13C-NMR (151 MHz, CDCl3)
δ (ppm) = 118.74 (C-3’), 118.95 (C-2), 118.99 (C-5’), 120.06 (C-1’),
128.66 (C-4”), 129.65 (C-6’), 129.68 (C-5”), 132.89 (C-3”), 136.49 (C-4’), 138.00 (C-3), 140.30 (C-2”),
163.69 (C-2’), 193.27 (C-1).
3.2. Microorganisms
The studies were carried out on eight strains of yeasts of the species Rhodotorula rubra (KCh 4
and KCh 82), Rhodotorula marina (KCh 77), Rhodotorula glutinis (KCh 242), Yarrowia lipolytica (KCh
71), Candida viswanathii (KCh 120), Saccharomyces cerevisiae (KCh 464), and Candida parapsilosis (KCh
909), which have already been described [26,33], and come from the collection of the Department of
Chemistry of Wrocław University of Environmental and Life Sciences, Poland. All the strains were
cultivated on a Sabouraud agar consisting of aminobac (5 g), glucose (40 g) and agar (15 g) dissolved
in 1 L of distilled water and pH 5.5 and stored in a fridge at 4 ◦C.
3.3. Analysis
Initial tests were carried out using TLC plates (SiO2, DC Alufolien Kieselgel 60 F254 (0.2 mm thick),
Merck, Darmstadt, Germany). The product separation on a plate in cyclohexane is Ethyl acetate eluent
(9:1 v/v). The product was observed (without additional visualization) under the UV lamp for the
wavelength of 254 nm.
3.4. Gas Chromatography (GC)
GC analysis were performed using an Agilent 7890A gas chromatograph, equipped with
a flame ionization detector (FID) (Agilent, Santa Clara, CA, USA). The capillary column DB-5HT
(
30 m × 0.25 mm × 0.10 µm) was used to determine the composition of the product mixtures.
A temperature program was applied as follows: 80–300 ◦C, the temperature on the detector: 300 ◦C,
injection 1 l, flow 1 mL/min, flow H2: 35 mL/min, air flow; 300 mL/min, time of analysis; 18.67 min.
The retention time of the substrate —9,6 min, product retention time—8,5 min. The retention times
and were recorded as 10,8 min, and 9,7 min,
µ
1
2
for compounds having the thiophene substituent
respectively (Supplementary Materials).
3
4
3.5. NMR Analysis
NMR analysis was performed using a DRX 600 MHz Bruker spectrometer (Bruker, Billerica,
MA, USA). The prepared samples were dissolved in deuterated chloroform CDCL3. The analyses
performed, included 1H-NMR, 13C-NMR, HMBC (two-dimensional analysis) HMQC (heteronuclear
correlation) and COSY (correlation spectroscopy) (Supplementary Materials).

Molecules 2019, 24, 3185
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3.6. Screening
Erlenmeyer flasks with a capacity of 300 mL were used for biotransformation; each contained
100 ml of the culture medium – Sabouraud (3% glucose, 1% peptone). The transplanted microorganisms
were incubated for three days at 24 ◦C on a rotary shaker (144 rpm). After this time, the substrates
1
and
3
were added in an amount of 10 mg, dissolved in 1 mL of DMSO (dimethyl sulfoxide). Samples were
collected after 1, 3, 6, 12 hours and 1, 3, and 7 day of incubation. After this time, samples were extracted
with ethyl acetate, dried with anhydrous magnesium sulfate (MgSO4), and analyzed by TLC and GC.
3.7. Semi-Preparative Scale
Semi-preparative biotransformations were performed in 2 L Erlenmeyer flasks, each containing
500 mL of culture medium (3% glucose, 1% peptone). The transferred microorganisms were incubated
◦
for three days at 24 C on a rotary shaker. After this time, the substrate was added in 100 mg,
dissolved in 2 mL of DMSO. After ten days, the product in the mixture was isolated by triple extraction
with ethyl acetate (3 extractions of 300 mL). After drying with anhydrous magnesium sulfate, and
concentrating the sample by using a rotary evaporator, the obtained product was analyzed by GC
and NMR.
The biotransformation product was separated on preparative TLC plates (1000 µm, silica gel
plates (Anatech, Gehrden, Germany) with cyclohexane: Ethyl acetate eluent (9:1 v/v). After separation,
the product was scraped from the plate, extracted twice from the gel with ethyl acetate, dried with
magnesium sulfate and concentrated by a rotary vacuum evaporator. A one-day transformation
of 3-(2”-furyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (
(colorless oil) of 3-(2”-furyl)-1-(2’-hydroxyphenyl)-propan-1-one (
3-(2”-furyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one ( ) (100mg) in the same strain culture yielded 76 mg
1
) (100mg) in Y. lipolytica KCh 71 gave 72 mg
2). One-day transformation of
3
(pale yellow oil) of 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propan-1-one (
then analyzed by NMR spectroscopy.
4). The resulting products were
4. Conclusions
There is known catalytic activity of ene reductases present in many yeast species, consisting in
the hydrogenation of chalcone to dihydrochalcone. In this study, 3-(2”-furyl)- (1) and
3-(2”-thienyl)-1-(2’-hydroxyphenyl)-prop-2-en-1-one (3) were biotransformed using eight yeast strains.
The purpose was to test the microorganisms’ ability to selectively hydrogenate the double bond in
chalcone containing heteroatom, in one of the rings, and select the most efficiently transforming
strain. A significant factor in this type of reaction is not only its duration and the efficiency
with which the substrate is transformed, but also the lack of by-products. Out of the available
microorganisms, the fastest conversions of substrate 1 to the expected product were observed in
Saccharomyces cerevisiae KCh 464 and Y. lipolytica KCh 71 strains. Product 2 was observed following
the first hour of biotransformation (constitutive enzyme), and the conversion efficiency >99% was
recorded as early as six hours after the start of the biotransformation. High conversion of substrate
1, higher than 90% after 24 hours of biotransformation, was also observed in the R. rubra KCh
4 culture. However, when analyzing the composition of the reaction mixture in twelve hours,
the observed substrate conversion did not exceed 10%. This biotransformation pattern indicates that it
is probably the result of substrate-induced ene reductases. Comparing these data to the conversion of
2-hydroxychalcone to dihydrochalcone, as previously described in [32], where the above-mentioned
strains were also tested, we observed that the presence of the heteroatom in the B-ring practically does
not affect the transformation efficiency—after 72 hours in the case of KCh 71 and KCh 464 strains;
however, it is much more efficient in the case of the strain KCh 4. Substrate 3, which is composed
of a sulfur atom in ring B, was analogously converted by the tested biocatalysts in the same way
as substrate 1. Substrate 3 was most effectively transformed by Y. lipolytica KCh 71 strain culture.

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During this biotransformation, 3-(2”-thienyl)-1-(2’-hydroxyphenyl)-propan-1-one (4) was obtained in
99% yield after the third hour of transformation.
Supplementary Materials: Supplementary materials are available online.
Author Contributions: Formal analysis, M.L., M.K. and T.J.; investigation, M.L. and M.K.; methodology, M.L., T.J.
and E.K.-S.; project administration, M.L. and T.J.; resources, T.J.; supervision, T.J. and E.K.; visualization, M.L.;
writing-original draft, M.L.; writing-review and editing, M.L. and T.J.
Funding: This research received no external funding
Conflicts of Interest: The authors declare no conflict of interest.
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2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
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