SHORT REPORT
Rec. Nat. Prod. 10:3 (2016) 392-396
Chemical Constituents of Jacaranda oxyphylla and their
Acetylcholinesterase Inhibitory and Antimicrobial Activities
Vinicius Viana Pereira1,2*, Roqueline Rodrigues Silva3,
Lucienir Pains Duarte1 and Jacqueline Aparecida Takahashi1
1
Chemistry Department, Universidade Federal de Minas Gerais, 31270-901, Belo Horizonte, Brazil
2
Faculty of Pharmacy, Universidade Federal de Minas Gerais, 31270-901, Belo Horizonte, Brazil
3
Chemistry Department, Universidade Federal dos Vales do Jequitinhonha e Mucuri,
39100-000, Diamantina, Brazil
(Received February 8, 2015; Revised March 04, 2015; Accepted March 05, 2015)
Abstract: This study evaluated chemical composition of Jacaranda oxyphylla, acetylcholinesterase inhibitory and
antimicrobial activities of the isolated compounds. Phytochemical investigation of leaves extract yielded three
classes of substances: fatty compounds, sterols and triterpenes. Butyl hexadecanoate (1), fatty alcohol (2), 2-(4hydroxyphenyl)ethyl triacontanoate (3), β-sitosterol (4), sitosterol-3-O-β-D-glucoside (5), 6'-palmitoyl-sitosterol-3O-β-D-glucoside (6), oleanolic acid (7), ursolic acid (8) and corosolic acid (9) were obtained from n-hexane,
CHCl3 and EtOH extracts of J. oxyphylla. It was found a pronounced acetylcholinesterase inhibitory activity for
the fatty compounds 1-3 and sterols 5 and 6, with values between 60 to 77%. Substances 7-9 presented a high
antibacterial action against Bacillus cereus and Salmonella typhimurium, with values of growth inhibition in the
range of 84 to 90%.
Keywords: Bignoniaceae; Jacaranda oxyphylla; acetylcholinesterase inhibition; antibacterial activity. © 2015
ACG Publications. All rights reserved.
1. Plant Source
Jacaranda oxyphylla Cham. is found in the Brazilian Cerrado region, popularly known as
“caroba-de-São-Paulo” and it is used in folk medicine to treat microbial infections1,2. Due to some
similarities, J. oxyphylla has been previously identified as a variety of the medicinal plant J. caroba.
However, these species can be differentiated by analysis of their respective leaflets, which are ellipticlanceolate with 7-9 secondary veins in J. oxyphylla3.
*
Corresponding author: E- Mail: vncsviana@yahoo.com.br; Phone +55-31-3409-6846
The article was published by Academy of Chemistry of Globe Publications
www.acgpubs.org/RNP © Published 10/12/2015 EISSN: 1307-6167
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Pereira et al., Rec. Nat. Prod. (2016) 10:3 392-396
The aerial parts of J. oxyphylla were collected in São João da Chapada, near Diamantina city in
July 2012. Plant material was identified by Dr. L. H. Y. Kamino (Institute of Biological Science,
Universidade Federal de Minas Gerais, Brazil) and a voucher specimen (No. 170.970) was deposited in
BHCB Herbarium of the same university.
2. Previous Studies
Fatty materials have been found in the extracts of Jacaranda species4. For example, 2-(4hydroxyphenyl)ethyl triacontanoate (3) was isolated from the stem of J. filicifolia and showed inhibitory
activity against the 5-lipoxygenase enzyme5.
Phytosterols are metabolites widespread in plant species and can be found as free alcohols,
esterified to fatty acids or as glycosides6. β-sitosterol (4) was previously isolated from J. filicifolia5, J.
mimosifolia7 and J. caroba; sitosterol-3-O-β-D-glucoside (5) was identified from the stem bark of J.
mimosifolia8. Plant sterols have drawn attention due to biological activities featured by them. There is
evidence that some phytosterols are effective in preventing cardiovascular diseases9.
Acid triterpenes of different skeleton have been isolated from Jacaranda species4. Oleanolic acid
(7) was previously isolated from J. mimosifolia7 and J. caroba; ursolic acid (8) was identified in J.
filicifolia5, J. caroba, J. copaia10 and J. decurrens11; corosolic acid (9) was previously isolated from J.
caucana12. Several biological effects are associated with triterpenes, such as antitumor, antiinflammatory, antimicrobial and anti-HIV activities13,14.
3. Present Study
After drying at room temperature, leaves and twigs of J. oxyphylla were separated and
powdered. Dried leaves of J. oxyphylla (1.2 kg) were extracted successively with n-hexane, CHCl3 and
EtOH by maceration. Extracts were prepared at room temperature, followed by filtration. n-Hexane,
CHCl3 and EtOH extracts were concentrated under vacuum using a rotary evaporator to afford crude
extracts as follows: n-hexane extract (13 g), CHCl3 extract (56 g) and EtOH extract (215 g).
Part of the crude n-hexane, CHCl3, and EtOH (10, 20, and 20 g, respectively) extracts were
submitted to silica gel 60 column chromatography (n-hexane, CHCl3, EtOAc and MeOH as eluents, in
order of increasing polarity). Fractions of 150 mL were collected and concentrated under vacuum in a
rotary evaporator. After thin layer chromatography analysis, similar fractions were pooled in groups.
Successive column chromatography purifications and recrystallizations were used for isolation and final
purification of compounds 1-9, that belong to different classes of phytochemicals (Figure 1). The
purified compounds were characterized using 1D and 2D NMR techniques, UV spectroscopy, mass
spectrometry analysis and comparison with previously reported spectral data15-20.
From the n-hexane extract, it was identified butyl hexadecanoate (1) (eluting with n-hexaneCHCl3 7:3; 87 mg), fatty alcohol (2) (eluted with n-hexane-CHCl3 1:1 and recrystallized with n-hexane;
68 mg), β-sitosterol (4) (eluting with CHCl3-EtOAc from 9:1 to 7:3; 94 mg) and oleanolic acid (7)
(eluted with CHCl3-EtOAc from 9:1 to 0:1 and recrystallized with EtOH; 11 mg). From the CHCl 3
extract, there were isolated the following compounds: 2-(4-hydroxyphenyl)ethyl triacontanoate (3)
(eluted with CHCl3-EtOAc 9:1 and recrystallized with n-hexane; 14 mg), β-sitosterol (4) (eluting with
CHCl3-EtOAc 9:1; 9 mg), sitosterol-3-O-β-D-glucoside (5) (eluted with EtOAc-MeOH from 9:1 to 1:1
and recrystallized with (CH3)2CO; 7 mg), 6'-palmitoyl-sitosterol-3-O-β-D-glucoside (6) (eluting with
CHCl3-EtOAc 1:9; 50 mg), oleanolic acid (7) and ursolic acid (8). Oleanolic and ursolic acids have very
similar structures and the separation of these compounds is not easy. The CHCl 3 extract was subjected
to column chromatography over silica gel and eluted gradient with CHCl3-EtOAc from 1:0 to 3:7. It was
obtained a fraction with oleanolic acid (156 mg), an intermediate fraction with oleanolic and ursolic
acids mixed (570 mg) and another fraction with ursolic acid (601 mg). These substances were purified
by recrystallization with EtOH. From the EtOH extract, ursolic acid (8) (740 mg) and corosolic acid (9)
(624 mg) were isolated over silica gel eluting with CHCl3-EtOAc 3:2 to EtOAc-MeOH 1:1. These acid
triterpenes were purified by recrystallization with EtOH.
394
Chemical constituents of Jacaranda oxyphylla
(1)
(7)
(4) R= OH
(2)
(5) R=
(6) R=
(8) R= H
(9) R= OH
(3)
Figure 1. Structures of compounds 1-9 isolated from leaves of J. oxyphylla.
Acetylcholinesterase inhibition: Chemical constituents (1-9) isolated from J. oxyphylla leaves were
screened on a quantitative assay for measuring acetylcholinesterase inhibition (iAChE), based on
Ellman’s method21. It was found a potential acetylcholinesterase inhibitory activity for the fatty
materials 1-3 and sterols 5 and 6 with values between 60.9 to 77.7% of inhibition, as presented in Table
1. It was observed that the presence of glycosides in the structure of sterols 5 and 6 makes these
compounds at least eight times more potent if compared to their precursor, compound 4. The hydroxyl
moieties present in 5 and 6 could be involved in hydrogen bonding with the amino acid residues of the
active site of the acetylcholinesterase enzyme22.
Table 1. In vitro antiacetylcholinesterase activity (iAChE) and growth inhibition of microorganism
induced by compounds 1-9 isolated of J. oxyphylla.
Compound
iAChE*
Microorganism growth inhibition*
S. aureus
E. coli
S. typhimurium
ATCC 29212 ATCC 25922 ATCC 14028
17.8 ± 0.7
0
9.1 ± 0.8
1
60.9 ± 1.4
B. cereus
ATCC 1778
23.6 ± 1.4
C. albicans
ATCC 18804
5.8 ± 1.1
2
77.7 ± 1.2
23.3 ± 1.6
31.1 ± 0.8
11.8 ± 0.8
11.6 ± 1.2
14.7 ± 1.6
3
75.4 ± 1.3
28.1 ± 1.5
38.2 ± 1.2
16.8 ± 0.7
18.2 ± 1.4
35.7 ± 1.5
4
8.0 ± 1.0
19.0 ± 1.0
26.7 ± 1.3
10.6 ± 0.8
8.3 ± 0.9
37.1 ± 1.4
5
65.0 ± 1.3
20.9 ± 2.0
33.1 ± 0.9
17.7 ± 0.8
18.3 ± 1.3
31.0 ± 1.4
6
72.8 ± 1.5
35.3 ± 1.3
12.9 ± 0.8
0
45.8 ± 1.5
24.0 ± 1.2
7
0
90.3 ± 1.4
30.4 ± 1.0
49.6 ± 0.9
87.1 ± 1.1
36.6 ± 1.6
8
0
88.7 ± 1.5
27.6 ± 0.8
40.1 ± 0.7
85.8 ± 0.9
33.7 ± 1.1
9
0
85.8 ± 1.1
27.7 ± 0.9
41.1 ± 0.8
84.5 ± 0.9
52.1 ± 1.4
Standard**
87.8 ± 0.7
98.5 ± 0.5
99.3 ± 0.4
99.4 ± 0.3
97.8 ± 1.1
97.9 ± 1.1
*Results are mean values of quintuplicate assays ± standard deviation (expressed as % inhibition); **eserine for
acetylcholinesterase, ampicillin for bacteria and nystatin for yeast; compounds were assayed in concentration of 100 µg mL-1.
Antimicrobial screening: Compounds 1-9 obtained from leaves of the J. oxyphylla were subjected to
antimicrobial assay by broth microdilution method23. Gram-positive bacteria Bacillus cereus and
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Pereira et al., Rec. Nat. Prod. (2016) 10:3 392-396
Staphylococcus aureus, Gram-negative bacteria Escherichia coli and Salmonella typhimurium and the
yeast Candida albicans were tested. Substances 7-9 presented a high antibacterial action against B.
cereus and S. typhimurim, with values of growth inhibition in the range of 84.5 to 90.3%. Moreover,
triterpene 9 presented a moderate activity against C. albicans (52.1%). The overall results of the
antimicrobial assay are shown in Table 1.
This study reported the isolation of nine compounds from the leaves of J. oxyphylla, a species
without chemical and biological studies in the literature. β-sitosterol and its glycosides derivatives
(compounds 4-6) were the phytosterols obtained and the triterpenoid acids isolated were olean-12-ene or
urs-12-ene derivatives (compounds 7-9). It was obtained a high quantity of ursolic acid (3.7% of EtOH
extract), corosolic acid (3.1% of EtOH extract) and oleanolic acid (0.8% of CHCl3 extract). Thus, J.
oxyphylla revealed to be a natural source of these triterpenes, which exhibited a high antibacterial
activity. This is the first report on the isolation of compounds 2 and 6 in Bignoniaceae family. These
fatty compounds have potential inhibitory activity towards acetylcholinesterase and could be useful as
lead for developing alternative drugs to the treatment of Alzheimer's disease.
Acknowledgements
Authors are grateful to Conselho Nacional de Desenvolvimento Científico e Tecnológico,
Coordenação de Aperfeiçoamento de Pessoal de Nível Superior and Fundação de Amparo à Pesquisa do
estado de Minas Gerais for financial support.
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