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showed by methanol extract of E. calophrys stems
(Sahidin et al., 2018). Other studies revealed the
potency of E. littoralis rhizomes and E. maingayi
leaves as antibacterial agents (Chiang et al, 2010).
Moreover, essential oil of Etlingera fenzlii (Kurz) K.
Schaum was safe for repellent source (Sudhakaran et
al, 2016).
According to the above information, the
chemistry and pharmacology aspects of E. elatior
fruits have not been reported. Hence, the present
work will facilitate and report the isolation and
identification of chemical compounds from the
methanol extract of E. elatior fruits, as well as their
antibacterial activity.
2 MATERIALS AND METHODS
2.1 General Procedures
Instruments were used Cary Varian 100 Conc UV
spectrophotometer, PerkinElmer Spectrum One FT-
IR spectrophotometer, and JEOL ECP 500 NMR
spectrometer (500 MHz for
1
H and 125 MHz for
13
C). Chromatography techniques were performed
using Kieselgel 60 F
254
0,25 mm, silica gel 60 GF
254
,
and silica 60 G (Merck, Darmstadt, Germany). TLC
plates were derivatised using a cerium sulphate
reagent (Merck, Darmstadt, Germany). DPPH (2,2-
diphenyl-1-picrylhydrazyl) was purchased from
Merck (Darmstadt, Germany).
2.2 Sample
Fruits of Etlingera elatior were collected from the
Wolasi Forest, South Konawe, South East Sulawesi,
in November 2016 with No of Specimen EST02.
The plant specimen was identified and stored in the
Herbarium Bogoriense, Indonesia.
2.3 Extraction and Isolation
The dried powdered fruits of E. elatior (2.1 kg) was
macerated with methanol (MeOH, 3 x 5.0 L, 24 h
each time) at room temperature and yielded a dried
methanol extract as dark green gum (80 g). This
extract was further fractionated using a silica gel
VLC (10 x 5 cm, 150 g), eluted with n-hexane–ethyl
acetate (from 9:1 to 0:10) followed by pure MeOH,
and gave 5 main fractions (F1-F5) with weight of
1.3, 4.1, 7.3, 6.2, and 29.6 g, respectively. Main
fraction F3 was re-fractionated using a silica gel
VLC (10 x 5 cm, 150 g) and gradiently eluted with
n-hexane–ethyl acetate (from 7:3 to 0:10) and
MeOH as mobile phases, to yield subfractions F31
(0.1 g), F32 (0.7 g), F33 (0.7 g), and F34 (4.4 g).
Subfraction F32 was chromatographed using a silica
gel RC with chloroform–MeOH (95:5) and pure
MeOH as mobile phases, to produce pure
compound 1 (0.03 g). Furthermore, subfraction F33
was further purified using the same method as
compound 1 purification to get compound 2 (0.08 g).
2.4 Antibacterial Activity
The antibacterial assay was determined against
Bacillus subtilis FNCC 0060, Escherichia coli
ATCC 35218, Pseudomonas aeruginosa ATCC
27853, Salmonella enterica ATCC 14028,
Staphylococcus aureus ATCC 25923, and
Streptococcus mutans ATCC 25175. The
antibacterial test was conducted by the agar dilution
method using the general procedure outlined by
Thakurta (Sahidin et al, 2017). The cultural
concentration of bacteria was (B. subtilis = 2.0 x 10
8
cfu/mL, E. coli= 4.2 x 10
8
cfu/mL, P. aeruginosa =
1.2 x 10
8
cfu/mL, S. enterica = 2.0 x 10
8
cfu/mL, S.
aureus= 3.2 x 10
7
cfu/mL and S. mutans= 1.2 x 10
7
cfu/mL).
3 RESULTS
3.1 Physicochemical Property and
Spectroscopic Data of The Isolated
Compounds From E. Elatior Fruits
Two compounds (1–2) were successfully isolated
and identified from the methanol extract of E. elatior
fruits. Structures of these compounds were
determined based on their physicochemical property
and spectroscopic spectra of IR and NMR. These
data were also compared with the same data reported
in the previous studies.
Vanilic Acid (1); a white powder. Spectra of
1
H
NMR (500 MHz, CDCl
3
) δ
H
(ppm): 10.87 (br, s),
8.40 (1H, s), 7.61 (1H, dd, 8.4, 1.9), 7.58 (1H, d,
1.,7), 6.93 (1H, d, 8.2), and 3.92 (3H, s). Spectra of
13
C NMR (125 MHz, CDCl
3
) δ
C
(ppm): 166.6 (C-7),
151.2 (C-4), 147.2 (C-3), 124.0 (C-6), 122.0 (C-1),
112.6 (C-5) and 55.4 (C-8).
p-Hydroxybenzoic acid (2); white amorphous
powder. Spectra of
1
H NMR (500 MHz, CDCl
3
) δ
H
(ppm): 9.43 (1H, s), 7.91 (2H, d, J = 8.6 Hz, H-2/H-
6), 6.92 (2H, d, J = 8.4 Hz, H-3/H-5). Spectra of
13
C
NMR (125 MHz, CDCl
3
) δ
C
(ppm): 166.7 (C-7),
BROMO 2018 - Bromo Conference, Symposium on Natural Products and Biodiversity
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