A straightforward synthesis of natural oxygenated ent-kaurenoic acid derivatives

Article abstract of DOI:10.1080/00397911.2020.1821225

The paper presents the synthesis of some natural diterpenoids of ent-kauranic structure, starting from ent-kaur-16-en-19-oic acid, a natural diterpenoid readily available from the dried wastes of sunflower (Helianthus annuus L.). The reported ent-16α,17-dihydroxykauran-19-oic, ent-16α-hydroxy-17-acetoxykauran-19-oic and ent-16α,17-diacetoxykauran-19-oic acids have been previously isolated from different plant sources and showed diverse biological activities, including anti-HIV, anti-cancer and anti-Alzheimer properties. Their chemical synthesis has not been disclosed so far. Additional cytotoxicity studies of the obtained compounds revealed a remarkable cell proliferative effect of parent ent-kaur-16-en-19-oic acid and two of its synthetic derivatives, which is in line with recent pioneering studies in this area.

Full text of DOI:10.1080/00397911.2020.1821225

Synthetic Communications
An International Journal for Rapid Communication of Synthetic Organic
Chemistry
ISSN: (Print) (Online) Journal homepage: https://www.tandfonline.com/loi/lsyc20
A straightforward synthesis of natural oxygenated
ent-kaurenoic acid derivatives
Olga Morarescu , Marina Grinco , Veaceslav Kulciţki , Alic Barba , Olga
Garbuz , Valentin Gudumac , Aurelian Gulea & Nicon Ungur
To cite this article: Olga Morarescu , Marina Grinco , Veaceslav Kulciţki , Alic Barba , Olga
Garbuz , Valentin Gudumac , Aurelian Gulea & Nicon Ungur (2020): A straightforward synthesis
of natural oxygenated ent-kaurenoic acid derivatives, Synthetic Communications, DOI:
10.1080/00397911.2020.1821225
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SYNTHETIC COMMUNICATIONSV
https://doi.org/10.1080/00397911.2020.1821225
A straightforward synthesis of natural oxygenated
ent-kaurenoic acid derivatives
Olga Morarescua, Marina Grincoa, Veaceslav Kulcit¸kia , Alic Barbaa,
Olga Garbuzb,c, Valentin Gudumacd, Aurelian Guleab, and Nicon Ungura
aLaboratory of Chemistry of Natural and Biologically Active Compounds, Institute of Chemistry MECR,
b
ꢀ
ꢀ
Chis¸inau, Republic of Moldova; Department of Chemistry, Moldova State University MECR, Chis¸inau,
c
Republic of Moldova; Laboratory of Molecular Systematics and Phylogeny, Institute of Zoology MECR,
d
ꢀ
Chis¸inau, Republic of Moldova; Laboratory of Biochemistry, Nicolae Testemitanu State University of
ꢀ
Medicine and Pharmacy, Chis¸inau, Republic of Moldova
ABSTRACT
ARTICLE HISTORY
Received 5 August 2020
The paper presents the synthesis of some natural diterpenoids of
ent-kauranic structure, starting from ent-kaur-16-en-19-oic acid, a nat-
ural diterpenoid readily available from the dried wastes of sunflower
(Helianthus annuus L.). The reported ent-16a,17-dihydroxykauran-19-
oic, ent-16a-hydroxy-17-acetoxykauran-19-oic and ent-16a,17-diace-
toxykauran-19-oic acids have been previously isolated from different
plant sources and showed diverse biological activities, including anti-
HIV, anti-cancer and anti-Alzheimer properties. Their chemical syn-
thesis has not been disclosed so far. Additional cytotoxicity studies
of the obtained compounds revealed a remarkable cell proliferative
effect of parent ent-kaur-16-en-19-oic acid and two of its synthetic
derivatives, which is in line with recent pioneering studies in this
area.
KEYWORDS
Allylic isomerization;
bioactivity; cell proliferation;
dihydroxilation; epoxidation;
ent-kaurane; ent-16a-
hydroxy-17-acetoxykauran-
19-oic acid; ent-kaur-16-en-
19-oic acid; ent-16a17-
dihydroxylkauran-19-oic
acid; ent-16; a,17-
diacetoxykauran-19-oic acid;
ent-16a17-epoxykauran-19-
oic acid; ent-17-oxo-16R-
kauran-19-oic acid; ent-17-
hydroxykaur-15-en-19-oic
acid; natural diterpenoids
GRAPHICAL ABSTRACT
H
H
16
H
OR1
OR2
OH
[O], H+
[O]
H
H
H
H
CO2H
H
CO2H
H
CO2H
4 R1 = R2 = H
1
6
9
R1 = OAc; R2 = H
10 R1 = R2 = OAc
Introduction
Ent-kauranic diterpenoids have been isolated from different plants and possess a broad
spectrum of biological activities.[1,2] The interest toward this group of diterpenoids is
still growing, provided the relevant application potential foreseen for a whole array of
such compounds.[3]
One of the most widespread diterpenoids of ent-kauranic structure is the ent-kaur-16-
en-19-oic acid (1). It was found among major secondary metabolites from different parts
CONTACT Nicon Ungur
nicon.ungur@gmail.com
Laboratory of Chemistry of Natural and Biologically Active
ꢀ
Compounds, Institute of Chemistry MECR, 3, Academiei str., Chisinau, MD 2028, Republic of Moldova.
,
Supplemental data for this article can be accessed on the publisher’s website.
ß 2020 Taylor & Francis Group, LLC

2
O. MORARESCU ET AL.
of numerous plants, including Viguiera sp.[4], Annona sp.[5], Wedelia sp.[6], Copaifera
sp.[7], Mikania sp.[8] One of the most available sources of acid (1) is Helianthus annuus
L. (sunflower)[9], which represents a global industrial crop.
Despite its availability, acid 1 was not actively explored for the synthesis of other nat-
ural kauranes. Most synthetic works relate on the chemical modifications of carboxylic
functional group within diverse SAR studies.[10] In the meanwhile, elaboration of syn-
thetic methods toward other functionalized derivatives of compound 1 represents a sure
path to access bioactive natural diterpenoids, which could be investigated under broader
medicinal chemistry studies.
Basing on our previous results on the isolation of 1 from whole wastes of H. annuus
L.[11] harvesting, we expanded our research to the use of this compound for the synthe-
sis of its functionalized derivatives.
Results and discussion
The natural ent-kaur-16-en-19-oic acid (1) possesses an exocyclic double bond, which
can be used as a helpful synthetic handle for the generation of different oxygenated
products. Surprisingly, this prospect was not explored on the free acid 1 but on its pro-
tected derivatives instead, comprising mainly methyl ester or different amides.[12]
Despite the fact that protected carboxyl group in 1 was considered as a pre-requisite of
enhanced biological activity, there are still a lot of interesting ent-kauranes with the free
angular carboxyl group in cycle A. We decided to investigate the reactivity of free acid
1 under different reaction settings, oriented toward exocyclic bond oxygenation. This is
the most straightforward way to access related natural ent-kauranoic acids functional-
ized at C16 and C17 atoms.
For practical reasons, the double bond oxygenation in acid 1 can be more conveni-
ently performed on epoxidation with a peracid. The strong steric hindrance around the
b-face of the molecule could provide a ground for oxygen addition facial selectivity.
The use of freshly prepared monoperphthalic acid (MPPA)[13] resulted in a good con-
1
version to the expected epoxides 2 and 3 with a satisfactory selectivity (ꢀ2:9, H-NMR
and GC-MS data). This mixture of epoxides was separated chromatographically on a sil-
ica gel column on eluting with mixtures of petroleum ether and ethyl acetate of increas-
ing polarities. The minor (ꢀ16%) epoxide 2 was eluted first. Its structure was
demonstrated on the basis of spectral data (IR, 1 D and 2 D NMR spectra). The major
(ꢀ44%) epoxide 3 was successively eluted and its structure was demonstrated basing on
the spectral data and also on comparing with the previously published data.[14] It is
noteworthy mentioning that a partial reactivity of the oxyranic cycle was observed dur-
ing separation on silica gel.
Application of m-CPBA as an alternative epoxidation agent led to a better conversion
(ꢀ93%) with a total selectivity toward the a-epoxide 3. The reaction was performed in
chloroform according to the described methodology (Scheme 1).[14]
The obtained major epoxide 3 was envisioned as a bridge toward other more func-
tionalized products. In particular, routine epoxide opening under acidic conditions was
investigated as a route to the natural dihydroxylated ent-kaurane 4. This transformation
was attempted on treatment of epoxide 3 with 70% perchloric acid in a DMSO-water

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SYNTHETIC COMMUNICATIONSV
3
12
H
H
16
OH
H
16
OAc
11
20
13
14
17
1
4
9
16
2
3
OR
OH
10
H 8
e
f
5
H
H
15
7
~94 %
~71%
6
H
H
H
18
19CO2H
g
CO2H
9 R = H
10 R = Ac
CO2H
4
1
~95%
a or b
H
16
H
16
H
16
H
OH
CHO
O
O
H
c or d
+
H
H
H
H
H
+
H
CO2H
H
H
CO2H
CO2H
CO2R
5
R = H
6
2
3 R = H
7 R = Me
8
R = Me
Scheme 1. Reagens and conditions: (a) MPPA, Et2O, rt, 2 h, ꢀ90%, (2)/(3) ¼ 2:9; (b) m-CPBA CHCl3,
rt, 24 h, ꢀ93% (3); (c) HClO4 (70%), DMSO/H2O, 45 ꢂC, 4 h, ꢀ26% (5), ꢀ44% (6); (d) FSO3H, i-PrNO2-
CH2Cl2, ꢁ60 ꢂC, 15 min, ꢀ75% (5), ꢀ2% (6). (e) OsO4, K3Fe(CN)6, t-BuOH, H2O, rt, 48 h, ꢀ71%; (f) Ac2O,
Py, rt, 12 h, ꢀ94%; (g) Ac2O, Py, 4-DMAP, rt, 24 h, ꢀ95%.
mixture. Usual workup and chromatographic separation of reaction products resulted in
isolation of the ent-17-oxo-16R-kauran-19-oic acid (5), along with ent-17-hydroxykaur-
15-en-19-oic acid (6) with a combined yield of ꢀ70% in a ꢀ2:3 relative ratio. It was
obvious that the reaction path included competitive oxirane isomerization and opening.
No diol product was formed and this result can be accounted for strong sterical effects
of the ent-kauranic skeleton. Our attempt to change the acid catalyst and use a super-
acid (FSO3H, i-PrNO2-CH2Cl2, ꢁ78 ꢂC, 15 min) led to a more selective isomerization to
the ent-17-oxo-16R-kauran-19-oic acid (5), formed with a ꢀ75% yield. The allylic alco-
hol 6 was minor under these conditions (ꢀ2%).
The prevailing acid 5 is a new synthetic compound, which was not reported from
any natural source yet. Its methyl ester 7 has been described previously, as a
rearrangement product of epoxide 8.[15] Literature data on the epoxide 8 rearrange-
ment are quite contradictory. The authors[15a] describe the conversion of 8 under
the action of different Lewis acids at room temperature into a ꢀ1:1 mixture of epi-
meric aldehydes at C16. The same Lewis acid-induced rearrangement of 8 shown in
[15b]
ꢁ
Hueso-Falcon et al.
relates on a totally selective rearrangement, but the resulting
aldehyde is provided with the opposite stereochemistry as described in Batista
et al.[15a] We believe that our conditions, involving low temperature superacidic
treatment of epoxides contribute to a high selectivity of the rearrangement pro-
cess.[16] From the mechanistic point of view, this rearrangement follows a usual
pathway, having as a key step the 1,2-hydride shift, which is totally selective at low
temperature due to the inability of the bulky OH-group to adopt a sterically hin-
dered b-orientation (structure A, Scheme 2).
The structure of ent-17-oxo-16R-kauran-19-oic acid (5) is unambiguously confirmed,
basing on extensive 1 D and 2 D NMR experiments. The IR spectrum shows absorption
bands specific to aldehyde and carboxylic groups (1691 cmꢁ1 and 2737cmꢁ1 respectively).
1
The H-NMR spectrum contains singlets at d 1.23 and 0.92, c corresponding to geminal

4
O. MORARESCU ET AL.
H
H
H
H
H
H
CHO
H
OH
H
16
H
+H+
O
H
H
H
H
~H
+
+
-H+
OH
5
3
A
Scheme 2. Proposed mechanism of epoxide 3 rearrangement.
C18 equatorial and C20 axial methyl groups of the ent-kaurane backbone. The triplet sig-
nal of one methyne C16 proton at 2.58ppm and the doublet signal of the aldehydic C17
13
proton at 9.64–9.65 ppm are clearly observed. The ‘-NMR spectrum shows the signals
of quaternary carbons at C4, C8 and C10 at 43.7, 45.1 and 39.6 ppm respectively, along
with carboxylic and aldehydic C19 and C17 carbons found at 184.0 and 203.7 ppm respect-
ively. The signals of trisubstituted C5, C9, C13 and C16 carbons are presented at 56.9, 55.2,
37.8 and 53.6 ppm. The relative stereochemistry of the CHO-group was established basing
on NOE correlation between protons H11$H16$H15$H9 and an obvious sign of a lack
of interaction between H16$H14 protons. The a-orientation of the CHO group was con-
firmed by comparison of the splitting pattern and chemical shift of the C17 proton with
known data from similar compounds.[15]
The structure of ent-17-hydroxy-kaur-15-en-19-oic acid 6 was also confirmed on the
1
basis of its spectral (IR, H- and 13C-NMR) data, corresponding exactly those reported
previously for the natural analogue, isolated from roots of Inula japonica Thunb[17a]
and Aralia cordata Thunb.[17b] It is noteworthy mentioning that the acid 6 has been
isolated from different parts of other plant sources, including Espeletiopsis,[18]
Helianthus,[9f,19] and showed anti-Alzheimer activity.[3a]
The dihydroxilation of acid 1 was successfully achieved on catalytic osmilation
with OsO4-K3Fe(CN)6 according to the known method.[20] The desired ent-16a,17-
dihydroxykauran-19-oic acid (4) was obtained with an yield of ꢀ71% after chroma-
1
tographic separation on a silica gel column. All physicochemical data (IR, H- and
13C-NMR) of the synthetic 4 matched the literature values.[5c,5d,21]
The ent-16a,17-dihydroxykauran-19-oic acid (4) was isolated previously from some
species of HeIianthus sp.[9d,22], Wedelia calycina[23] and Siegesbeckia pubescens medicinal
plants,[24] leaves, bark and green fruits of Xylopia frutescens,[25] and from the fruits of
some Annona sp.[5,22] Investigation of its biological activity resulted in the discovery of
anti-HIV,[5c,22] anti-cancer[5d] and anti-Alzheimer effect induced by pure 4.[3a] An inter-
esting property of this compound was described in a pioneering work of Sung et al.[26]
The authors demonstrated a remarkable regeneration of keratinocyte stem/progenitor
cells in epidermal tissue induced by natural 4 isolated from Siegesbeckia pubescens
leaves. This finding opens promising prospects for natural ent-kauranes to be applied in
surgery and tissue engineering. Some more recent data indicate on a significant inhib-
ition of lipopolysaccharide-induced nitric oxide production in microglia BV2 type
cells.[27] This activity connects directly to a potential neuroinflamatory modulator prop-
erties of the acid 4. Surprisingly, but there is only one report on the synthesis of 4 on
the oxidation of 1 with dimethyldioxyrane.[28] It was formed along with other oxygen-
ated derivatives in a yield of ꢀ18%.

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5
Acetylation of the acid 4 under standard condition with acetic anhydride in pyridine led
to the formation of the ent-16a-hydroxy-17-acetoxykauran-19-oic acid (9) with a ꢀ94%
yield. The structure of synthetic compound 9 was established basing on its spectral data
(IR, 1H- and 13C-NMR), that correspond to those reported previously for the natural ana-
logue.[29a] It was isolated from the roots of Annona glabra Linn.[28a] and aerial part of
Inula japonica Thunb plant.[29b] The acid 9 showed a significant ability to inhibit nitric
oxide production in RAW264.7 macrofage cells with IC50 value of 14.3 mg/mL.[29b]
The acetylation of compound 9 under more forced conditions in the presence of 4-DMAP
as an acyl transfer catalyst provided after 24 hours of stirring at room temperature the ent-
16a,17-diacetoxykauran-19-oic acid (10) with a yield of ꢀ95%. The structure and stereo-
1
chemistry of the formed product 10 is demonstrated by its spectral data (IR, H- and 13C-
NMR). In particular, the relative stereochemistry was clearly demonstrated basing on NOESY
correlations, showing strong interaction between C17 (4.93, 4.42 ppm), C15 (1.92, 1.67ppm)
and C11 (1.61, 1.49 ppm) protons, along with weak C17 (4.93, 4.42 ppm)—C13 (2.47 ppm) and
absence of C17 (4.93, 4.42 ppm)—C14 (1.93, 1.42 ppm) protons interactions. The HSQC—
HMBC spectra allowed us to adjust the chemical shift assignment published previously for
the natural diacetate 10, isolated from the stems of Annona senegalensis plant.[30]
The natural compound 10 exhibited cytotoxic selectivity for PC-3 (prostate cancer)
cells but with moderate potencies (ED50 17–18 mg/mL).[30] The chemical synthesis of
both products 9 and 10 was not reported previously.
In vitro evaluation of the antiproliferative and cytotoxic activity
The antiproliferative activity of the tested compounds 1, 3, 9 and 10 on cervical epithe-
lial cell of line HeLa, human epithelial pancreatic adenocarcinoma cell of line BxPC-3,
human muscle rhabdomyosarcoma cell of line RD and Madin Darby Canine Kidney
epithelial normal cell line MDCK were determined, using the resazurin cytotoxic assay,
which allows one to measure the number of viable cells.[31] To estimate the results on
the cytostatic effect of the cell of lines, the tested compounds were compared to doxoru-
bicine (DOXO) as the referent control, which is used in the clinical management of a
wide range of cancers.
Study of antiproliferative activity showed that there is concentration dependence
between inhibitory effects of the tested compounds at the micromolar range. The inhibi-
tory effect was displayed in a dose-dependent manner. Comparative study of antiproli-
ferative activity of the tested compounds and the referent controls DOXO at
concentration 10 mM in regard to lines cancer cells HeLa, BxPC-3, RD and normal cells
MDCK are shown in Figure 1.
The inhibitory rates of the tested compounds on cancer cells HeLa, BxPC-3, RD were
lower, than DOXO. Thus, 1, 3, 9 and 10 exhibited cytotoxic activity against tested can-
cer cell lines, with IC50 values ꢃ100 mM. The IC50 values of lines HeLa, BxPC-3, RD are
2.8 5.7 mM, 4.0 7.9 mM, 1.31 4.9 mM for DOXO, respectively. The tested compounds
1, 3, 9 and 10 at concentration 10 mM inhibit proliferation of cancer cells lines vary in
the range 24.7–13.8%, 20.4–1.4%, 16.1–5.8% and 23.2–2.6%, respectively.
The positive control DOXO demonstrated cytostatic effect against MDCK cell lines
with IC50 value of 1.5 0.1 mM. The tested compound 3 at concentration 10 mM inhibits

6
O. MORARESCU ET AL.
90
81.4
80
70
63.9
62.4
58.3
60
DOXO (10 µM)
3 (10 µM)
50
40
1 (10 µM)
24.7
23.2
30
20.4
18.6
10 (10 µM)
9 (10 µM)
16.1
20
15.3
13.8
7.8
8.1
6.2
5.8
10
2.6
1.4
0
HELA
BxPC-3
RD
MDCK
−1
−2.2
−10
−20
−5.2
Figure 1. Comparative study of antiproliferative activity of the tested compounds 1, 3, 9 and 10.
the formation and growth of MDCK cell lines by 8.1%. As can be seen from Figure 1,
the tested compounds 1, 9 and 10 at 10 mM show inhibitory activity on cancer cells pro-
liferation, but, on the contrary, enhance the proliferative effect on normal cells MDCK,
which indicates its nontoxic properties with respect to the MDCK line. These results
denote a clear perspective of kauranes 1, 9 and 10 as potential tissue regeneration
agents, in line with the demonstrated properties of the diol 4.[26]
Conclusion
The synthesis of several natural diterpenoids belonging to the ent-kaurane series has been
demonstrated starting from ent-kaur-16-en-19-oic acid (1) on oxidative functionalization
at C16 and C17 carbon atoms. The obtained ent-16a,17-dihydroxykauran-19-oic (4), ent-
16a-hydroxy-17-acetoxykauran-19-oic (9), ent-16a,17-diacetoxykauran-19-oic (1) and ent-
17-hydroxykaur-15-en-19-oic (6) acids are biologically active natural compounds showing
anti-HIV, anti-cancer and anti-Alzheimer activity. Basing on the elaborated synthetic
scheme, larger amounts of these compounds can be synthesized for the expansion of bio-
logical activity studies to include unexplored fields. In particular, compounds 1, 3 and 9
show a proliferative effect on normal MDCK cells, which indicates their nontoxic proper-
ties regarding the MDCK line and a cellular growth promotion potential. These data are
convergent to the pioneering studies on the regeneration of keratinocyte stem/progenitor
cells in epidermal tissue under the action of similar natural ent-kauranes.
Experimental section
General
Melting points were measured with a Boethius heating stage. Optical rotations: Jasco-
DIP-370 polarimeter; 5-cm cell; in CHCl3. IR Spectra: Spectrum-100 FT-IR spectropho-
1
tometer (Perkin ꢁ Elmer), with the universal ATR sampling accessory; ꢀ in cmꢁ1. H-

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SYNTHETIC COMMUNICATIONSV
7
and 13C-NMR Spectra: Bruker-Avance-III spectrometer (400.13 and 100.61 MHz); in
CDCl3; d in ppm rel. to CHCl3 as internal standard (d(H) 7.26 and d(C) 77.0), J in Hz.
GC/MS: Agilent-7890A chromatograph; quadrupole MS detector MSD 5975 C; HP-5ms
capillary column (30 m/0.25 mm). The carbon and hydrogen contents of compounds
were determined by standard microanalysis on Vario-EL-III-CHNOS Elemental
Analyzer. Commercial Merck silica gel 60 (70–230 mesh ASTM) was used for flash
chromatography and Merck pre-coated silica gel plates were used for TLC. The chroma-
tograms were sprayed with 0.1% solution of cerium (IV) sulfate in 2 N sulfuric acid,
and heated at 80 ꢂC for 5 min to detect the spots. Treatment of reaction mixtures in
organic solvents included the extraction by diethyl ether, washing of the extract with
water up to neutral reaction, drying over anhydrous Na2SO4, filtering, and solvent
removal in vacuum.
Isomerization of ent-16a,17-epoxykauran-19-oic acid (3)
(a) To a solution of ent-16a,17-epoxykauran-19-oic acid (3) (270 mg, 0.807 mmol) in
DMSO (9.0 mL) and H2O (4.5mL) was added HClO4 (70% solution, 1.34mL,
ꢀ9.30mmol). The reaction mixture was stirred for 4 h at 45ꢂC. Then the H2O (10.0 mL)
was added and after usual work-up the extract was dried and filtered. The solvent was
removed under reduced pressure. The residue (268 mg) was purified by silica gel (6 g)
column chromatography (petroleum ether—ethyl acetate, gradient elution) to aftord, in
order of increasing polarity, products 5 (53 mg, ꢀ20% yield) and 6 (118 mg, ꢀ44% yield),
which showed spectral data identical with those of described allylic alcohol.[17a]
(b) A solution of 3 (43 mg, 0.13 mmol) in 2-nitropropane (0.5 mL) and CH2Cl2 (1.0 mL)
was chilled to ꢁ60 ꢂC and treated with a solution of FSO3H (65 mg, 0.65 mmol) in 2-nitro-
propane (0.5 mL). After 15 min of stirring, quenching with Et3N-hexane (1:1, 2.0 mL) and
usual workup, the extract was dried and filtered. The solvent was removed under reduced
pressure on a rotary evaporator. According to NMR data the residue (39 mg) represents a
mixture (ꢀ97:3) of aldehyde 5 and alcohol 6. The residue was purified by silica gel (1.0 g)
column chromatography (petroleum ether—ethyl acetate, 9:1) to give 32 mg (ꢀ75% yield)
of ent-17-oxo-16R-kauran-19-oic acid (5) as a white crystalline solid. Mp: 196–198 ꢂC;
20
1
½aꢄD ꢁ126.1 (c 2.2, CHCl3); IR (ꢀ, cmꢁ1): 2737, 1691, 1451, 1266, 1242; H-NMR (CDCl3,
400.13 MHz, dH ppm): 0.75–0.89 (2, m), 0.92 (3, s, Me-20), 0.95–1.10 (3, m), 1.23
(3, s, Me-19), 1.39–1.62 (75, m), 1.63–1.91 (7, m), 2.11–2.19 (1, m), 2.50–2.55
(1, m), 2.58 (1, t, J¼ 7.0 Hz, C-16), 9.65 (1, d, J¼ 1.6 Hz, CHO); 13C-NMR (CDCl3,
100.61 MHz, dC ppm): 15.5 (q, C-20), 18.8 (t, C-11), 19.0 (t, C-2), 22.3 (t, C-6), 28.9 (q,
C-18), 30.9 (t, C-12), 37.7 (t, C-14), 37.8 (t, C-3), 37.8 (d, C-13), 39.6 (s, C-10), 40.1 (t, C-
15), 40.7 (t, C-1), 41.0 (t, C-7), 43.7 (s, C-4), 45.1 (s, C-8), 53.6 (d, C-16), 55.2 (d, C-9),
56.9 (d, C-5), 184.0 (s, C-19), 203.7 (s, C-17); GC-MS: 318, 300, 275, 261, 243, 215, 136,
123, 109, 91; Anal. Calc. for C20H30O3: C, 75.43; H, 9.50; Found: C, 75.45; H, 9.51.
1
Full experimental detail, including original NMR spectra, assignment of H and 13C
NMR spectra for compounds 2, 5 and 10 and biological assay results. This material can
be found via the “Supplementary Content” section of this article’s webpage.

8
O. MORARESCU ET AL.
Funding
The financial support from National Agency for Research and Development (ANCD) of the
Republic of Moldova [Project No 20.80009.8007.03] is gratefully acknowledged.
ORCID
Veaceslav Kulci¸tki
http://orcid.org/0000-0002-9363-1615
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