Biotransformation of testosterone by Cladosporium sphaerospermum

Article abstract of DOI:10.1080/10242422.2019.1583747

Incubation of testosterone 1 with Cladosporium sphaerospermum MRC 70266 afforded six metabolites and two of these metabolites, 6β,16β,17β-trihydroxyandrost-4-en-3-one 6 and 6β,12β,17β-trihydroxyandrost-4-en-3-one 7, were determined as new compounds. The fungus mainly hydroxylated testosterone 1 at C-6β, accompanied by some minor hydroxylations at C-7β, C-12β, C-15α and C-16β. A minor oxidation at C-17 and a minor 5α-reduction were also observed.

Full text of DOI:10.1080/10242422.2019.1583747

Biocatalysis and Biotransformation
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Biotransformation of testosterone by
Cladosporium sphaerospermum
Kudret Yildirim, Ali Kuru & Şengül Yılmaz
To cite this article: Kudret Yildirim, Ali Kuru & Şengül Yılmaz (2019): Biotransformation of
testosterone by Cladosporiumꢀsphaerospermum, Biocatalysis and Biotransformation, DOI:
10.1080/10242422.2019.1583747
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BIOCATALYSIS AND BIOTRANSFORMATION
https://doi.org/10.1080/10242422.2019.1583747
RESEARCH ARTICLE
Biotransformation of testosterone by Cladosporium sphaerospermum
€
and S¸engul Yılmaz
Kudret Yildirim , Ali Kuru
Department of Chemistry, Faculty of Arts and Sciences, Sakarya University, Sakarya, Turkey
ABSTRACT
ARTICLE HISTORY
Received 27 December 2018
Accepted 8 February 2019
Incubation of testosterone 1 with Cladosporium sphaerospermum MRC 70266 afforded six metab-
olites and two of these metabolites, 6b,16b,17b-trihydroxyandrost-4-en-3-one 6 and 6b,12b,17b-
trihydroxyandrost-4-en-3-one 7, were determined as new compounds. The fungus mainly
hydroxylated testosterone 1 at C-6b, accompanied by some minor hydroxylations at C-7b, C-
12b, C-15a and C-16b. A minor oxidation at C-17 and a minor 5a-reduction were also observed.
KEYWORDS
Testosterone; biotransform-
ation; Cladosporium
sphaerospermum
Introduction
Materials and methods
Microbial steroid biotransformations have found
worldwide application for the preparation of more
valuable steroidal drugs and hormones due to their
high regio- and stereoselectivities. There are still enor-
mous efforts to perform more effective microbial ster-
oid biotransformations and to detect new useful
microorganisms and reactions (Donova and Egorova
2012; Nassiri-Koopaei and Faramarzi 2015).
Cladosporium, belonging to the Cladosporiaceae
family of the Ascomycota, is a large genus (Sandoval-
Denis et al. 2016). Cladosporium species are cosmopol-
itan in distribution and usually isolated from air, food,
plants, paint and textiles (Bensch et al. 2012; Sandoval-
Denis et al. 2016). These species are common endo-
phytes, plant pathogens and even hyperparasites of
other fungi (Sandoval-Denis et al. 2016). Some
Cladosporium species are also considered pathogenic to
humans and animals (Sandoval-Denis et al. 2015).
Cladosporium sphaerospermum is a halotelorant or
osmotelorant fungus widely distributed around the
world (Zalar et al. 2007). It is frequently isolated from
indoor and outdoor locations (Aihara et al. 2001; Park
et al. 2004), humans (Yano et al. 2003) and plants
(Pereira et al. 2002).
Instrumental methods
1H NMR spectra were recorded in deuteriochloroform
with tetramethylsilane as an internal standard refer-
ence at 300 MHz with a Varian Mercury 300 spec-
trometer (Palo Alto, CA). 13C NMR spectra were
recorded in deuteriochloroform at 75 MHz with a
Varian Mercury 300 spectrometer. Chemical shifts are
given in ppm (d scale), and coupling constants (J)
are given in Hz. Infrared spectra were recorded using
a
Perkin Elmer Spectrum Two spectrometer
(Whaltam, MA). Elemental analysis was performed
using a Thermo Finnigan Flash EA 1112 elemental
analyser (Bremen, Germany). Melting points were
determined by an Electrothermal IA 9200 melting
point apparatus (Staffordshire, UK) and are uncor-
rected. Thin layer chromatography (TLC) was carried
out with 0.2 mm thick Merck Kieselgel 60 F254 TLC
plates (Istanbul, Turkey) using ethyl acetate/n-hexane
(1:1) as eluent. TLC plates were dipped into an ani-
ꢀ
saldehyde/H2SO4 reagent and heated to 120 C for 3
min in order to visualize the spots.
Chemicals
As far as steroid biotransformations by C. sphaero-
spermum are concerned, we have not found any litera-
ture work on steroids. In this work, testosterone 1 was
incubated with C. sphaerospermum MRC 70266 for 5 d
in order to investigate its metabolism.
Testosterone 1 was purchased from Sigma-Aldrich
(Istanbul, Turkey). Solvents were of analytical grade
and were purchased from Merck (Istanbul, Turkey).
Potato dextrose agar (PDA) and agar for PDA slopes
CONTACT Kudret Yildirim
kudrety@sakarya.edu.tr
Department of Chemistry, Faculty of Arts and Sciences, Sakarya University, Sakarya
54187, Turkey
Supplemental data for this article can be accessed here.
ß 2019 Informa UK Limited, trading as Taylor & Francis Group

2
K. YILDIRIM ET AL.
and ingredients for liquid medium were also pur-
chased from Merck (Istanbul, Turkey).
broth was then extracted three times with each 1 L of
ethyl acetate. The organic extract was dried over
anhydrous sodium sulphate, and the solvent evapo-
rated in vacuo to give a brown gum (1759 mg). The
steroids in brown gum were chromatographed on sil-
ica gel 60 (Merck 107734), eluting with increasing con-
centrations of ethyl acetate in n-hexane. Steroid
mixtures from the column were then separated by col-
umn chromatography on aluminium oxide 90 active
neutral (activity stage I, Merck 101077), using smaller
columns for 2–3 h. The purified steroids were crystal-
lized in appropriate solvents and identified by using
melting points and a combination of NMR, IR and
elemental analysis.
Micro-organism and fermentation conditions
Cladosporium sphaerospermum MRC 70266 was obtained
from TUBITAK, Marmara Research Center, Food Science
and Technology Research Institute, Culture Collection
Unit, Kocaeli, Turkey. Stock cultures were maintained at
ꢀ
on PDA slopes at 4 C.
The liquid medium for the C. sphaerospermum MRC
70266 was prepared by mixing glucose (20 g), pep-
tone (5 g) and yeast extract (5 g) in 1 L of distilled
water (Shebany 2012). The medium was evenly distrib-
uted among 10 culture flasks of 250 mL capacity (100
mL in each) and autoclaved for 20 min at 121 ꢀC.
Spores freshly obtained from PDA slopes were trans-
ferred aseptically into each flask containing sterile
medium in a biological safety cabinet. After cultivation
Biotransformation of testosterone 1 by C.
sphaerospermum MRC 70266
Incubation of testosterone 1 with C. sphaerospermum
MRC 70266 for 5 d afforded 6 metabolites 2–7
(Figure 1). Elution with ethyl acetate in n-hexane (2:3)
afforded the unreacted substrate (97 mg), which was
ꢀ
at 28 C for 3 d on a rotary shaker (160 rpm), testos-
terone 1 (1 g) dissolved in 10 ml of DMF was evenly
distributed aseptically among the flasks. The biotrans-
formation of the substrate was carried out in 10 flasks
for 5 d under the same conditions. Biotransformation
experiment was run with a control flask containing
non-inoculated sterile medium and the substrate.
After 5 d of incubation, the control was harvested and
analysed by TLC. No metabolites were detected in
the control.
1
identified by comparison of its H and 13C NMR spec-
tra with those of an authentic sample.
Elution with ethyl acetate in n-hexane (1:1) afforded
6b-hydroxyandrost-4-en-3,17-dione 2 (84 mg, 8%);
ꢀ
crystallized from acetone as prisms; mp 189–190 C, lit
(Hanson et al. 1996); mp 190–191 ꢀC; IR ꢀmax 3415,
1740, 1670 cmÀ1; EA: calculated for C19H26O3: C 75.46,
1
H 8.67; found: C 75.32, H 8.53%; H NMR (CDCl3): dH
0.91 (3H, s, 18-H), 1.38 (3H, s, 19-H), 4.36 (1H, brs, 6a-
H), 5.80 (1H, s, 4-H); 13C NMR (CDCl3): see Table 1.
Elution with ethyl acetate in n-hexane (3:2) afforded
6b,17b-dihydroxyandrost-4-en-3-one 3 (422 mg, 40%);
crystallized from ethyl acetate as prisms; mp 212–213
Isolation and identification of metabolites
After incubation, the fungal mycelium was separated
from the broth by filtration under vacuum, and the
mycelium was rinsed with ethyl acetate (500 ml). The
Figure 1. Biotransformation of testosterone 1 by C. sphaerospermum MRC 70266.

BIOCATALYSIS AND BIOTRANSFORMATION
3
Table 1. 13C NMR data for testosterone
metabolites.
1
and its
200–201 ꢀC; IR ꢀmax 3445, 1680 cmÀ1; EA: calculated
for C19H28O4: C 71.22, H 8.81; found: C 71.10, H 8.73%;
1H NMR (CDCl3): dH 0.87 (3H, s, 18-H), 1.39 (3H, s, 19-
H), 3.50 (1H, dd, J = 5.0, 11.0Hz, 12a-H), 3.89 (1H, t, J =
8.5 Hz, 17a-H), 4.35 (1H, brs, 6a-H), 5.81 (1H, s, 4-H);
13C NMR (CDCl3): see Table 1.
C atom
1
2
3
4
5
6
7
1
2
35.48
33.74
36.97
34.09
36.37
34.19
36.93
34.24
35.68
33.89
37.04
34.19
35.02
34.13
3
199.64 200.33 200.41 214.07 199.61 200.47 200.26
4
5
123.87 126.34 126..32
171.52 168.06 168.32
34.61 123.76 126.38 126.53
39.91 171.07 168.14 167.67
6
7
8
9
10
11
12
13
14
15
16
17
18
19
32.63
31.33
35.58
53.69
38.45
20.10
36.21
42.61
50.24
23.14
29.51
72.58
37.17
29.34
53.57
38.00
20.19
31.19
47.57
50.81
21.64
35.71
72.96
37.09
29.75
53.64
38.01
20.56
38.01
42.88
50.44
23.25
30.42
81.64
11.07
19.51
29.96
69.63
37.81
59.24
36.50
20.26
42.25
43.29
51.29
23.04
29.44
81.12
10.96
22.98
32.72
32.14
35.29
53.81
38.63
20.50
36.56
44.31
58.43
72.46
42.57
78.70
12.56
17.48
72.90
38.04
29.17
53.81
38.15
20.27
36.98
42.43
46.94
34.91
69.85
80.62
11.88
19.52
72.81
37.43
28.84
52.46
37.95
29.33
78.99
46.98
48.51
23.07
30.01
81.87
6.02
Results
The first metabolite was identified as 6b-hydroxyan-
drost-4-en-3,17-dione 2. NMR spectra of 2 had new
resonances at dH 4.36 ppm (1H, brs) and dC 72.58 ppm
(see Supplemental data), which were typical for the
presence of a 6b-hydroxyl group (Hanson et al. 1996).
The 13C NMR spectrum of 2 lacked the C-17 resonance
of 1 at dC 81.25 ppm and had a new carbon atom res-
onance at dC 220.62 ppm, suggesting that an oxida-
tion had taken place at C-17.
81.25 220.62
10.93
17.17
13.71
19.46
19.44
ꢀC, lit (Hanson et al. 1996); mp 215–220 ꢀC; IR ꢀmax
3405, 1650, 1620 cmÀ1; EA: calculated for C19H28O3: C
The second metabolite was identified as 6b,17b-
1
1
dihydroxyandrost-4-en-3-one 3. The H NMR spectrum
74.96, H 9.27; found: C 74.82, H 9.13%; H NMR (CDCl3):
of 3 showed characteristic resonances at dH 3.65 ppm
(1H, t, J = 8.5 Hz) and dH 4.36 ppm (1H, brs), indicating
the presence of 17b- and 6b-hydroxyl groups, respect-
ively (Hanson et al. 1996). The 13C NMR spectrum of 3
had resonances at dC 72.96 and dC 81.64 ppm, which
were in agreement with the presence of 7b- and 17b-
hydroxyl groups, respectively.
dH 0.82 (3H, s, 18-H), 1.38 (3H, s, 19-H), 3.65 (1H, t, J =
8.5 Hz, 17a-H), 4.36 (1H, brs, 6a-H), 5.80 (1H, s, 4-H); 13C
NMR (CDCl3): see Table 1.
Elution with ethyl acetate in n-hexane (7:3) afforded
7b,17b-dihydroxyandrostan-3-one 4 (75 mg, 7%); crys-
ꢀ
tallized from ethyl acetate as needles; mp 202–203 C
ꢀ
lit (Farooq and Tahara, 2000); mp 196–197 C; IR ꢀmax
3325, 1710 cmÀ1; EA: calculated for C19H30O3: C 74.47,
The third metabolite was identified as 7b,17b-dihy-
droxyandrostan-3-one 4. The 1H NMR spectrum of 4
lacked the 4-H resonance of 1 at dH 5.74 ppm (1H, s),
suggesting the hydrogenation of the double bond in
ring A. The 13C NMR spectrum of 4 had two resonan-
ces at dC 69.93 ppm and dC 81.12 ppm, which were
typical for the presence of 7b- and 17b-hydroxyl
1
H 9.87; found: C 74.33, H 9.70%; H NMR (CDCl3): dH
0.74 (3H, s, 19-H), 0.83 (3H, s, 18-H), 3.65 (2H, m, 7a-H
and 17a-H); 13C NMR (CDCl3): see Table 1.
Elution with ethyl acetate in n-hexane (9:1) afforded
15a,17b-dihydroxyandrost-4-en-3-one 5 (53 mg,ꢀ 5%);
crystallized from acetone as needles; mp 96–97 C, lit
1
ꢀ
groups, respectively (Farooq and Tahara 2000). The H
(Peart et al. 2011); mp 93–94 C; IR ꢀmax 3400, 1660,
1650 cmÀ1; EA: calculated for C19H28O3: C 74.96, H
NMR spectrum of 4 showed two overlapping resonan-
ces at dH 3.65 ppm (2H, m), which were in agreement
with the presence of 7b- and 17b-hydroxyl groups.
The fourth metabolite was identified as 15a,17b-
dihydroxyandrost-4-en-3-one 5. The 13C NMR spectrum
of 5 showed two characteristic resonances at dH 3.89
ppm (1H, t, J = 8.5 Hz) and dH 4.11 ppm (1H, m), indi-
cating the presence of 17b- and 15a-hydroxyl groups,
respectively (Peart et al. 2011). The 13C NMR spectrum
of 5 showed two resonances at dC 72.46 and dC 78.70
ppm, further indicating the presence of 15a- and 17b-
hydroxyl groups, respectively.
The fifth metabolite was identified as 6b,16b,17b-
trihydroxyandrost-4-en-3-one 6. The 13C NMR spec-
trum of 6 had two resonances at dC 72.90 ppm and dC
80.62 ppm, which were characteristic (Hanson et al.
1996) for the presence of 6b- and 17b-hydroxyl
1
9.27; found: C 74.85, H 9.17%; H NMR (CDCl3): dH 0.80
(3H, s, 18-H), 1.20 (3H, s, 19-H), 3.89 (1H, t, J = 8.5 Hz,
17a-H), 4.11 (1H, m, 15b-H), 5.73 (1H, s, 4-H); 13C NMR
(CDCl3): see Table 1.
Elution with pure ethyl acetate afforded
6b,16b,17b-trihydroxyandrost-4-en-3-one
6 (34 mg,
3%); crystallized from ethyl acetate as cubes; mp
169–170 ꢀC; IR ꢀmax 3395, 1660 cmÀ1; EA: calculated
for C19H28O4: C 71.22, H 8.81; found: C 71.15, H 8.70%;
1H NMR (CDCl3): dH 0.88 (3H, s, 18-H), 1.39 (3H, s, 19-
H), 3.38 (1H, d, J = 7.3 Hz, 17a-H), 4.16 (1H, m, 16a-H),
4.34 (1H, brs, 6a-H), 5.81 (1H, s, 4-H); 13C NMR (CDCl3):
see Table 1.
Further elution with pure ethyl acetate afforded
6b,12b,17b-trihydroxyandrost-4-en-3-one
7 (23 mg,
2%); crystallized from ethyl acetate as needles; mp

4
K. YILDIRIM ET AL.
groups, respectively. The NMR spectra of 6 showed
new resonances at dH 4.16 ppm (1H, m) and dC 69.85
ppm, indicating the presence of another hydroxyl
group. The characteristic triplet of 17a-H (Bridgeman
et al. 1970) was observed as a doublet due to splitting
by a proton at C-16. The 13C NMR spectrum of 6
showed a downfield shift for C-15 (DdC 11.77 ppm)
whereas it showed a c-gauche upfield shift for C-14
(DdC 3.30 ppm), further indicating the presence of a
hydroxyl group at C-16. According to the literature,
the change in the characteristic multiplicity of 17a-H
resonance suggested the presence of a 16b-hydroxyl
minor oxidation at C-17 and a minor 5a-reduction
were also observed. In a recent work, however,
Cladosporium cladosporioides MRC 70282, another
Cladosporium isolate, metabolised 1 in a very different
way. This fungus only hydroxylated 1 at C-16b and
most of this new hydroxyl group was then oxidised to
a carbonyl group. A minor independent oxidation at
C-17 and a minor epimerisation at C-17 were also
observed (Yildirim et al. 2018a).
Reports of fungal 5a-reduction, oxidation at C-17
and hydroxylation of 1 at C-6b, C-15a and C-16b are
very common (Mahato and Garai 1997; Fernandes
et al. 2003; Bhatti and Khera 2012; Donova and
Egorova 2012; Nassiri-Koopaei and Faramarzi 2015).
However, fungal hydroxylations of 1 at C-7b and C-
12b are very rare (Tweit et al. 1962; Smith et al. 1990;
Farooq and Tahara 2000; Hunter et al. 2008; Yildirim
et al. 2018b). In a previous work, for example,
Botryosphaerica obtusa hydroxylated 1 at C-7b, C-11a,
C-12b and C-15a (Smith et al. 1990). In another work,
Botrytis cinerea hydroxylated 1 at C-7b and reduced its
C4–C5 double bond from the a-face (Farooq and
Tahara, 2000). Rhizomucor tauricus (Hunter et al. 2008)
hydroxylated 1 at C-6b and C-12b whilst a Penicillium
isolate (Tweit et al. 1962) hydroxylated 1 at C-1a, C-
6b, C-12b and C-15b. In a recent work, Ulocladium
chartarum MRC 72584 hydroxylated 1 at C-6b, C-7b, C-
12b and C-14a, accompanied by a 5a-reduction and
oxidations at C-6 and C-17 (Yildirim et al. 2018b).
However, in the present work, as stated above, C.
sphaerospermum MRC 70266 hydroxylated 1 at C-6b,
C-7b, C-12b, C-15a and C-16b, accompanied by a 5a-
reduction and an oxidation at C-17.
In short, we were able to see how testosterone 1
was metabolised by C. sphaerospermum MRC 70266. It
was shown that the fungus mainly hydroxylated 1 at
C-6b, accompanied by a minor oxidation at C-17, a
minor 5a-reduction and some minor hydroxylations at
C-7b, C-12b, C-15a and C-16b. In addition to this, two
of the metabolites, 6b,16b,17b-trihydroxyandrost-4-en-
3-one 6 and 6b,12b,17b-trihydroxyandrost-4-en-3-one 7,
were determined as new compounds. Our work on
biotransformations of some other steroids by C.
sphaerospermum MRC 70266 and some other fungi is
in progress.
group (Swizdor et al. 2017). The 13C NMR spectrum of
ꢀ
the metabolite exhibited resonances for 19 carbons
while its DEPT spectra revealed two methyl, six methy-
lene, seven methine and four quaternary carbons,
which were in accordance with the suggested structure
of the triol. 6b,16b,17b-Trihydroxyandrost-4-en-3-one 6
was determined as a new metabolite.
The sixth metabolite was identified as 6b,12b,17b-
trihydroxyandrost-4-en-3-one 7. The 13C NMR spec-
trum of 7 showed characteristic resonances at dC
72.81 ppm and dC 81.87 ppm, suggesting the pres-
ence of 6b- and 17b-hydroxyl (Hanson et al. 1996)
groups, respectively. The 13C NMR spectrum of 7 also
showed another characteristic resonance at dC 78.99
ppm, indicating the presence a 12b-hydroxyl group
(Hunter et al. 2008). The 13C NMR spectrum of 7
showed a downfield shift for C-11 (DdC 9.23 ppm)
whereas it showed a c-gauche upfield shift for C-14
(DdC 1.73 ppm), further indicating the presence of a
12b-hydroxyl group. The presence of two methyl, six
methylene, seven methine and four quaternary car-
bons in the DEPT spectra of 7 were in agreement with
the proposed structure of the triol. 6b,12b,17b-
Trihydroxyandrost-4-en-3-one 7 was determined as a
new metabolite.
Discussion
As can be seen from Table 2, C. sphaerospermum MRC
70266 hydroxylated testosterone 1 predominantly at
C-6b and some minor hydroxylations took place at C-
7b, C-12b, C-15a and C-16b. In addition to this, a
Table 2. Metabolite yields following chromatography.
Substrate
Metabolite
% Yield
Testosterone 1
6b-Hydroxyandrost-4-en-3,17-dione 2
6b,17b-Dihydroxyandrost-4-en-3-one 3
7b,17b-Dihydroxyandrostan-3-one 4
8
40
7
Acknowledgements
15a,17b-Dihydroxyandrost-4-en-3-one 5
6b,16b,17b-Trihydroxyandrost-4-en-3-one 6
6b,12b,17b-Trihydroxyandrost-4-en-3-one 7
5
3
2
The authors are grateful to Ras¸it Fikret Yılmaz and Fatih
Sonmez for running the NMR spectra.

BIOCATALYSIS AND BIOTRANSFORMATION
5
Park HG, Managbanag JR, Stamenova EK, Jong SC. 2004.
Comparative analysis of common indoor Cladosporium
species based on molecular data and conidial characters.
Mycotaxon. 89:441–451.
Peart PC, McCook KP, Russell FA, Reynolds WF, Reese PB.
2011. Hydroxylation of steroids by Fusarium oxysporum,
Exophiala jeanselmei and Ceratocystis paradoxa. Steroids.
76:1317–1330.
Pereira PT, De Carvalho MM, Girio FM, Roseiro JC, Amaral
CMT. 2002. Diversity of microfungi in the phylloplane of
plants growing in a Mediterranean ecosystem. J Basic
Microbiol. 42:396–407.
Disclosure statement
The authors report no declaration of interest. The authors
alone are responsible for the content and writing of
the paper.
ORCID
Kudret Yildirim
http://orcid.org/0000-0002-7889-5903
Ali Kuru http://orcid.org/0000-0002-8300-1572
€
S¸engul Yılmaz
http://orcid.org/0000-0001-5374-7448
Sandoval-Denis M, Sutton DA, Martin-Vicente A, Cano-Lira JF,
Wiederhold N, Guarro J, Gene J. 2015. Cladosporium spe-
cies recovered from clinical samples in the United States.
J Clin Microbiol. 53:2990–3000.
References
Aihara M, Tanaka T, Takatori K. 2001. Cladosporium as the
main fungal contaminant of locations in dwelling environ-
ments. Biocontrol Sci. 6:49–52.
Bensch K, Braun U, Groenewald JZ, Crous PW. 2012. The
genus Cladosporium. Stud Mycol. 72:1–401.
ꢀ
Sandoval-Denis M, Gene J, Sutton DA, Wiederhold NP,
Cano-Lira JF, Guarro J. 2016. New species of Cladosporium
associated with human and animal infections. Persoonia.
36:281–298.
Bhatti HN, Khera RA. 2012. Biological transformations of ster-
oidal compounds: a review. Steroids. 77:1267–1290.
Bridgeman JE, Cherry PC, Clegg AS, Evans JM, Jones ERH,
Kasal A, Kumar V, Meakins GD, Morisawa Y, Richards EE, et
al. 1970. Microbiological hydroxylation of steroids. Part I.
Proton magnetic resonance spectra of ketones, alcohols,
and acetates in the androstane, pregnane, and oestrane
series. J Chem Soc C. 250–257.
Shebany YM. 2012. Antimicrobial activity of endophytic fungi
isolated from Avicennia marina plant, Red Sea. Egypt J
Microbiol. 47:141–152.
Smith KE, Latif S, Kirk DN. 1990. Microbial transformations of
steroids-VI. Transformation of testosterone and andro-
stenedione by Botryosphaerica obtusa. J Steroid Biochem.
35:115–120.
ꢀ
Swizdor A, Panek A, Milecka-Tronina N. 2017. Hydroxylative
Donova MV, Egorova OV. 2012. Microbial steroid transforma-
tions: current state and prospects. Appl Microbiol
Biotechnol. 94:1423–1447.
activity of Aspergillus niger towards androst-4-ene and
androst-5-ene steroids. Steroids. 126:101–106.
Tweit RC, Dodson RM, Muir RD. 1962. Microbiological trans-
formations. XII. The substrate specificity of hydroxylations
by a Penicillium sp., A.T.C.C. 12,556. J Org Chem. 27:
3654–3658.
Farooq A, Tahara S. 2000. Biotransformation of testosterone
and pregnenolone catalyzed by the fungus Botrytis cin-
erea. J Nat Prod. 63:489–491.
Fernandes P, Cruz A, Angelova B, Pinheiro HM, Cabral JMS.
2003. Microbial conversion of steroid compounds: recent
developments. Enzyme Microbial Technol. 32:688–705.
Hanson JR, Nasir H, Parvez A. 1996. The hydroxylation of tes-
tosterone and some relatives by Cephalosporium aphidi-
cola. Phytochemistry. 42:411–415.
Hunter AC, Mills PW, Dedi C, Dodd HT. 2008. Predominant
allylic hydroxylation at carbons 6 and 7 of 4 and 5-ene
functionalized steroids by the thermophilic fungus
Rhizomucor tauricus IMI23312. J Steroid Biochem Mol Biol.
108:155–163.
Yano S, Koyabashi K, Kato K. 2003. Intrabronchial lesion due
to Cladosporium sphaerospermum in
asthmatic woman. Mycoses. 46:348–350.
a healthy, non-
Yildirim K, Kuru A, Yılmaz RF. 2018a. Microbial transformation
of some steroids by Cladosporium cladosporioides MRC
70282. J Chem Res. 42:408–411.
Yildirim K, Kuru A, Yılmaz S¸. 2018b. Biotransformation of tes-
tosterone by Ulocladium chartarum MRC 72584. J Chem
Res. 42:444–446.
Zalar P, de Hoog GS, Schroers H-J, Crous PW, Groenewald JZ,
Gunde-Cimerman N. 2007. Phylogeny and ecology of the
ubiquitous saprobe Cladosporium shaerospermum, with
descriptions of seven new species from hypersaline envi-
ronments. Stud Mycol. 58:157–183.
Mahato SB, Garai S. 1997. Advances in microbial steroid bio-
transformation. Steroids. 62:332–345.
Nassiri-Koopaei N, Faramarzi MA. 2015. Recent developments
in the fungal transformation of steroids. Biocatal
Biotransform. 33:1–28.