Home Antileishmanial, antibacterial and cytotoxicity activity of the extracts, fractions, and compounds from the fruits and stem bark extracts of Pentadesma butyracea Sabine
Article Publicly Available

Antileishmanial, antibacterial and cytotoxicity activity of the extracts, fractions, and compounds from the fruits and stem bark extracts of Pentadesma butyracea Sabine

  • Jean Koffi Garba , Ruland Tchuinkeu Nguengang , Gwladys Tatiana Youmbi , Joel Njopnu Menatche , Cyrille Armel Njanpa Ngansop , Jean Jules Kezetas Bankeu EMAIL logo , Jean Rodolphe Chouna ORCID logo , Fabrice Fekam Boyom ORCID logo , Norbert Sewald ORCID logo and Bruno Ndjakou Lenta EMAIL logo
Published/Copyright: October 21, 2021
Download Article (PDF)
Become an author with De Gruyter Brill
Author Information Explore this Subject
Zeitschrift für Naturforschung B
From the journal Zeitschrift für Naturforschung B Volume 77 Issue 1

Abstract

The search for antileishmanial plants used in traditional medicine led to the choice of CH2Cl2–MeOH (1:1) crude extract of the fruits and stem bark of Pentadesma butyracea Sabine (Clusiaceae) which displayed good activity in vitro against Leishmania donovani 1S (MHOM/SD/62/1S) promastigotes during preliminary screening with IC50 values 5.96 and 26.43 μg mL−1, respectively. The fractionation of both extracts using flash chromatography yielded active fractions with IC50 values ranging from 2.71 to 18.88 μg mL−1. Fourteen compounds (1–14) were isolated from the obtained fractions using successive column chromatographies and their structures were elucidated based on the analysis of their NMR and MS data. Daphnifolin (1), epicathechin (3), α-mangostin (9) and 9-hydroxycalabaxanthone (14) exhibited potent antileismanial activity against L. donovani 1S (MHOM/SD/62/1S) promastigotes with IC50 values of 2.01, 9.09, 3.37, and 6.87 μg mL−1, respectively and good selectivity towards Raw 264.7 macrophage cells (SI > 2.4). Extracts, fractions and some isolates were also assessed in vitro for their antibacterial activity against six bacterial strains [Salmonella typhi (CPC), Enterobacter cloacae (CPC), Pseudomonas aeruginosa HM801, Staphylococcus aureus ATCC 43300, Streptococcus pneumoniae ATCC 491619, Escherichia coli ATCC 25322] using serial microdilution method. Among the tested samples, the stem bark extract of P. butyracea as well as compounds 2 and 8 showed good to moderate activity against the aforementioned bacterial strains with MIC ≤ 250 μg mL−1.

Keywords: antibacterial; antileishmanial; clusiaceae; cytotoxicity; Pentadesma butyracea

1 Introduction

Leishmaniasis is a neglected tropical disease caused by an intracellular flagellate protozoan parasite belonging to the Leishmania genus. The disease is generally transmitted between man and animals during a blood meal by the phlebotome female sandfly [1]. About twenty different species of Leishmania were reported to be pathogenic to humans [2], including L. donovani, which is the deadliest visceral leishmaniasis. In 2017, 12 to 15 million peoples worldwide were infected and 350 million were at risk of acquiring the disease. An estimated 1.5 to 2 million of new cases and about 70000 related deaths are recorded each year [3]. Leishmaniasis is endemic in Africa, Asia, America, and the Mediterranean region. The three major clinical leishmaniasis forms occurring in humans are visceral (VL), cutaneous (CL) and mucocutaneous leishmaniasis (MCL) which differ in immunopathologies as well as in morbidity and mortality [4]. Cameroonian medicinal plants, from Clusiaceae family were reported as a source of potent antileismanial compounds [5]. This research was motivated by the fact that P. butyracea Sabine was reported to be used in traditional medicine for skin and hair care and in the manufacture of soap for healing qualities [6]. The macerated bark was also reported to be used in Gabon in lotions for the treatment of the parasitic diseases of the skin and as antidiarrhetic [7]. Different parts of this plant are used in tropical African medicine in the treatment of cough, fever, bronchitis, venereal diseases and viral infections [8]. To the best of our knowledge no antileishmanial potentials of P. butyracea is reported. In our continuing search for new antileishmanial agents from Clusiaceae family, we report herein the bioguided fractionation and isolation of secondary metabolites from the CH2Cl2–MeOH (1:1) stem bark and fruits extracts of P. butyracea, which displayed good leishmanicidal activity against L. donovani 1S (MHOM/SD/62/1S) promastigotes with IC50 values of 5.964 and 26.43 μg mL−1, respectively. In addition, they were assessed in vitro for their antibacterial activities against six bacterial strains including S. typhi (CPC), E. cloacae (CPC), P. aeruginosa HM801, S. aureus ATCC 43300, S. pneumoniae ATCC 491619, E. coli ATCC 25322.

2 Results and discussion

The CH2Cl2–MeOH (1:1) crude extracts of fruits and stem bark of P. butyracea exhibited a good in vitro antileishmanial activity against L. donovani 1S (MHOM/SD/62/1S) promastigotes with IC50 values 5.96 and 26.43 μg mL−1, respectively.

Figure 1: Structures of the isolated compounds.
Figure 1:

Structures of the isolated compounds.

The fruits extract of P. butyracea (PBF) was submitted to flash chromatography and afforded four main fractions labeled from PBFF1 to PBFF4. These fractions were assessed for their antileishmanial activity against L. donavani and exhibited good activity with IC50 values ranging from 12.94 to 25.97 μg mL−1 (Table 1), among which the ethyl acetate fraction (PBFF3) (IC50 = 12.94 μg mL−1) was the most active. It is noteworthy that fractionation has contributed to a decline in antileishmanial activity, suggesting that the activity of the crude extract could be due to the synergistic action of its constituents.

Table 1:

Antileishmanial activity and cytotoxicity of the extract, fractions and compounds from the fruits P. butyracea.

Extracts/compounds Antileishmanial activity IC50 ± SD (μg mL−1) Macrophages CC50 ± SD (μg mL−1) Selectivity index SI ± SD (=CC50/IC50)
PBF 5.96 ± 0.05d 398.70 ± 10.18d 66.89 ± 1.14g
PBFF1 17.37 ± 0.28i 106.35 ± 7.14c 6.11 ± 0.31b
PBFF2 25.97 ± 0.32l 397.25 ± 0.64d 15.29 ± 0.16c
PBFF3 12.94 ± 0.21g 485.55 ± 0.07f 37.52 ± 0.60f
PBFF4 18.88 ± 0.12j 465.50 ± 18.24e 24.33 ± 0.85d
1 2.01 ± 0.19b 66.68 ± 4.89b 33.21 ± 0.71e
2 16.59 ± 0.48h 22.18 ± 0.67a 1.33 ± 0.00a
3 9.09 ± 0.23e >100 ND
4 21.47 ± 0.25k >100 ND
5 27.00 ± 0.23m >100 ND
6 NA ND ND
7 11.04 ± 0.11f 75.50 ± 6.05b 6.83 ± 0.47b
8 >50 ND
9 3.37 ± 0.20c 80.79 ± 0.12b 24.02 ± 1.39d
Amphotericin B 0.22 ± 0.36a – ND

  1. ND, not determined; Data points are means from triplicate experiments. SD = Standard Deviation; Activity values were obtained from sigmoidal dose-response curves of concentration versus response. Along the columns values with different letter superscript are significantly different; Waller Ducan at p ≤ 0.05. PBF, P. butyracea fruit extract; PBFF1-4, P. butyracea fruit fractions. Bold values represent the numbering of compounds.

Further purification of the most active ethyl acetate fraction, PBFF3, led to the isolation of five compounds. Their structures were elucidated by comparison of their spectroscopic data with those from literature as daphnifolin (1) [9], norathyriol (2) [10], epicathechin (3) [11], methyl citrate (4) [12], and β-sitosterol-3-O-β-D-glucopyranoside (6) [13]. Furthermore, the chemical investigation of fraction PBFF2 afforded four compounds including β-sitosterol (5) [14], tovopyrifolin C (7) [15], cowagarcinone B (8) [16], and α-mangostin (9) [17] (Figure 1). Further investigation of PBFF4 and PBFF1 did not give a compound because of their complexity. The isolated compounds were assessed for their antileismanial activity against L. donovani 1S (MHOM/SD/62/1S) promastigotes and for their cytotoxicity towards Raw 264.7 macrophage cells (Table 1). Compounds 1 and 9 exhibited a good antileismanial activity against the parasite with IC50 values of 2.01 and 3.37 μg mL−1, respectively, and also showed a good selectivity towards (Raw 264.7 macrophage cells). Besides, the activity of compounds 1–3, 7, and 9 and those of fractions PBFF2 and PBFF3 could be due to antagonistic effects.

The CH2Cl2–MeOH (1:1) crude extract of stem bark of P. butyracea (PBB) was also submitted to flash chromatography to afford five fractions labeled from PBBF1 to PBBF5. These fractions exhibited good antileishmanial activity with IC50 values ranging from 2.71 to 13.56 μg mL−1 (Table 2). The fractions were then more active than the crude extract. The activity of the crude extract could be due to antagonistic effects of its constituents. The chemical investigation of fraction PBBF2 afforded eight compounds including daphnifolin (1) [9], β-sitosterol (5) [14], α-mangostin (9) [17], lupeol (10) [18], betulin (11) [18], tovophyllin A (12) [19], 1,3,7-trihydroxyxanthone (13) [20], and 9-hydroxycalabaxanthone (14) [10] (Figure 1). Fraction PBBF3 was further subjected to successive column chromatography and led to the isolation of three compounds including norathyriol (2) [10], epicatechin (3) [11] and β-sitosterol (5) [14]. Fractions PBBF1, PBBF4 and PBBF5 were not investigated because of their complexity. Compounds 10–14 were also assessed for their antileishmanial activity against L. donovani 1S (MHOM/SD/62/1S) promastigotes and for their cytotoxicity towards Raw 264.7 macrophage cells (Table 2). Among these isolates, compound 14 exhibited a good antileismanial activity against the parasite with IC50 value of 6.87 µg mL−1.

Table 2:

Antileishmanial activity and cytotoxicity of the extract, fractions and compounds from the stem bark of P. butyracea.

Extracts/compounds Antileishmanial activity (IC50 in μg mL−1) Macrophages CC50 ± SD (μg mL−1) Selectivity Index SI ± SD (=CC50/IC50)
PBB 26.43 ± 0.05k 148.81 ± 17.96e 5.62 ± 0.66c
PBBF1 7.91 ± 0.17f 106.35 ± 7.15d 13.45 ± 0.61d
PBBF2 12.56 ± 0.30i 48.775 ± 5.60b 3.77 ± 0.22b
PBBF3 2.71 ± 0.38c 75.58 ± 7.09c 28.01 ± 1.32f
PBBF4 12.75 ± 0.39i 405.80 ± 16.25g 31.82 ± 0.30g
PBBF5 10.88 ± 0.49h 305.00 ± 10.47f 28.04 ± 0.30f
10 >50 31.22 ± 1.64ab ND
11 >50 ND ND
12 >50 ND ND
13 >50 78.23 ± 7.81c ND
14 6.87 ± 0.33e 16.81 ± 0.67a 2.45 ± 0.02ab
Amphotericin B 0.22 ± 0.36a –

  1. ND, not determined; Data points are means from triplicate experiments. SD = Standard Deviation; Activity values were obtained from sigmoidal dose-response curves of concentration versus response. Along the columns values with different letter superscript are significantly different; Waller Ducan at p ≤ 0.05. PBB, P. butyracea stem bark extract; PBBF1-5, P. butyracea stem bark fractions. Bold values represent the numbering of compounds.

From the stem bark extract, fraction PBBF3 was the most active than the related compounds (2, 3, 5), while fraction PBBF2 was about four times less active than PBBF3, and some of its related highly active compounds (1, 9, 14). This may indicate that compounds from PBBF3 fraction have synergistic effect, while those from PBBF2 fraction have antagonist effect.

In conclusion, the fruit crude extract (PBF) was about four times more active than the stem bark extract (PBB). The fractionation of the PBF extract resulted in a decrease in the antileishmanial activity, while the fractionation of PBB extract led to an increase of activity, indicating the synergistic and antagonistic effect of fractions, respectively. The antileishmanial activity of daphnifolin (1) and α-mangostin (9) as well as 9-hydroxycalabaxanthone (14) corroborated with some previous results on xanthone derivative [21, 22]. Indeed, α-mangostin isolated from the fruits of Garcinia mangostana showed good activity against intracellular amastigotes of Leishmania infantum, with IC50 value 8 µM [23]. Daphnifolin (1) and α-mangostin (9), could be probably the main active principles isolated from both extracts. This result could justify the use of P. butyracea in the treatment of the parasitic diseases of the skin.

In addition, the fruits and stem bark extracts as well as some of the isolates were assessed for their antibacterial activity on six bacteria strains: E. coli ATCC 25322, S. pneumoniae ATCC 491619, P. aeruginosa HM801, S. typhi (CPC and CHU), E. cloacae (CPC), and S. aureus (CPC). PBB extract exhibited significant activity against the six strains, with MICs values ranging from 7.8 to 15.6 μg mL−1, while PBF extract was moderately active (Table 3). Compound 9 showed a good activity against the six strains with MICs ≤ 3.9 μg mL−1. These results are quite similar with those obtained by Koh and collaborator (2013) which showed that, α-mangostin isolated from the fruits of G. mangostana exhibited good activity against Gram-positive pathogens with MICs values between 0.78 and 1.56 μg mL−1 [24]. These evidences contribute to reinforce the knowledge on the potential of xanthone as potent antibacterial agents and thus, should draw awareness in the perspective of the search for new broad spectrum antibacterial agent from plant origin. In addition, it provides an insight that can justify the use of this plant in traditional medicine to treat skin and bacterial diseases [25, 26].

Table 3:

Antibacterial activity (MIC, μg mL−1) of extracts and compounds from the bark and fruits of P. butyracea.

Extracts/compounds Antibacterial activity (MIC in µg mL−1)
S. thyphi S. aureus E. cloacae P. aeruginosa S. pneumoniae E. coli
PBF – 125 500 500 500 500
PBB 7.8 15.6 7.8 7.8 31.2 15.6
1 – – 500 – – –
2 250 31.2 62.5 250 – 125
3 – 250 500 – – –
6 – 250 – 250 250 500
7 – – 250 – – –
8 125 15 125 250 125 125
9 <3.9 <3.9 3.9 3.9 3.9 3.9
Ciprofloxacin 0.03 0.15 0.06 0.07 0.03 0.03

  1. Salmonella typhi (CPC and CHU), Enterobacter cloacae (CPC), Pseudomonas aeruginosa HM801, Staphylococcus aureus (CPC), Streptococcus pneumoniae ATCC 491619; E coli ATCC 25322. CPC: “Centre Pasteurˮ of Cameroon. CHU: “Centre Hospitalier Universitaireˮ of Cameroon. Bold values represent the numbering of compounds.

3 Experimental

3.1 General experiment procedures

The 1H and 13C NMR spectra were recorded on Bruker DRX spectrometers at 500 and 125 MHz, respectively. Column chromatography (open column) was performed with silica gel Merck 60 (0.063–0.200 mm) and percolated aluminum backed silica gel 60 F254 sheets were used for TLC. Size exclusion CC was performed using Sephadex LH-20. The TLC spots were visualized under UV light (254 and 365 nm); H2SO4 conc. (10%) was used as spraying reagents.

3.2 Plant material and identification

The stem bark and fruits of P. butyracea were collected in October 2018 at Bazou, West Region of Cameroon. The plant material was identified at the National Herbarium of Cameroon, Yaoundé, by comparison with voucher specimens formerly kept under the registration number No 6861/SRF/Cam.

3.3 Extraction and isolation

The fruits and stem bark of P. butyracea were chopped into pieces, air dried under shade, and ground to give 4.8 and 3.5 kg of powder, respectively, and were separately macerated with (1:1) CH2Cl2-MeOH (3 × 5 L) twice for 48 h at room temperature (26 °C). The extracts of fruits and stem bark were freed from solvent under vacuum to yield 210.7 and 190.8 g, respectively. These extracts were submitted to bioguided fractionation towards L. donovani 1S (MHOM/SD/62/1S) promastigotes strain. The fruits crude extract (205.1 g) was subjected to flash chromatography over silica gel to afford four main fractions PBFF1 [n-hexane-EtOAc (1:0), 80.5 g], PBFF2 [n-hexane-EtOAc (1:1), 70.8 g], PBFF3 [n-hexane-EtOAc (0:1), 15.6 g], and PBFF4 [EtOAc-MeOH (4:1) to MeOH, 30.9 g]. The most active fraction PBFF3 was subjected to several column chromatography (CC) over silica gel and eluted with the mixtures of n-hexane-EtOAc (3:2–4:1) of increasing polarities and afforded compounds 1 (22.5 mg), 2 (6.5 mg), 3 (68.4 mg), 4 (420.0 mg), and 6 (5.2 mg). Fraction PBFF2 was also subjected to repeated CC over silica gel and eluted with the mixtures of n-hexane-EtOAc (9:1–8:2) to afford compounds 5 (9.5 mg), 7 (340.0 mg), 8 (5.4 mg), and 9 (14.2 mg).

The stem bark crude extract was submitted to flash column chromatography and afforded five fractions PBBF1 [n-hexane-EtOAc (1:0), 15.3 g), PBBF2 [n-hexane-EtOAc (1:1), 85.6 g], PBBF3 [n-hexane-EtOAc (0:1), 19.7 g], PBBF4 [EtOAc-MeOH (9:1), 19.5 g], and PBBF5 [EtOAc-MeOH (3:1) to MeOH (0:1), 30.3 g]. The most active fraction PBBF3 was submitted to CC over silica gel, and eluted with the mixtures of n-hexane-EtOAc (3:7–0:1) and afforded compounds 2 (25.6 mg), 3 (96.6 mg), and 6 (8.0 mg). Fraction PBBF2 was subjected to CC over silica gel and led to the isolation of compounds 10 (5.1 g), 9 (136.9 mg), 5 (41.1 mg), 11 (10.2 mg), 12 (4.2 mg), 1 (158.5 mg), 13 (3.8 mg), and 14 (25.0 mg).

3.3.1 Daphnifolin (1)

1H NMR (500 MHz, CD3OD-CDCl3, 25 °C, TMS): δ = 6.58 (s, 1 H, 2-H), 7.24 (dd, J = 7.8, 1.7 Hz, 1 H, 6-H), 7.20 (t, J = 7.8 Hz, 1 H, 7-H), 7.65 (dd, J = 7.8, 1.7 Hz, 1 H, 8-H), 3.91 (s, 3 H, 4-OMe). – 13C NMR (125 MHz, CD3OD-CDCl3): δ = 158.6 (C-1), 94.2 (C-2), 153.2 (C-3), 130.7 (C-4), 145.8 (C-4a), 154.1 (C-5), 120.1 (C-6), 123.6 (C-7), 115.2 (C-8), 120.8 (C-8a), 181.4 (C-9), 103.1 (C-9a), 145.4 (C-10a), 60.1 (OMe).

3.3.2 Norathyriol (2)

1H NMR (500 MHz, CD3OD, 25 °C, TMS): δ = 6.16 (d, J = 2.2 Hz, 1 H, 2-H), 6.31 (d, J = 2.2 Hz, 1 H, 4-H), 6.83 (s, 1 H, 5-H), 7.45 (s, 1 H, 8-H). – 13C NMR (125 MHz, CD3OD): δ = 163.0 (C-1), 97.3 (C-2), 164.9 (C-3), 93.4 (C-4), 158.0 (C-4a), 102.1 (C-5), 143.4 (C-6), 151.8 (C-7), 107.7 (C-8), 112.5 (C-8a), 179.7 (C-9), 101.9 (C-9a), 153.9 (C-10a).

3.3.3 Epicatechin (3)

1H NMR (500 MHz, CD3OD, 25 °C, TMS): δ = 4.83 (s, 1 H, 2-H), 4.19 (brs, 1 H, 3-H), 2.88 (dd, J = 16.8, 4.6 Hz, 1 H, 4a-H), 2.75 (dd, J = 16.8, 3.0 Hz, 1 H, 4b-H), 5.93 (d, J = 2.3 Hz, 1 H, 6-H), 5.96 (d, J = 2.3 Hz, 1 H, 8-H), 6.99 (d, J = 2.0 Hz, 1 H, 2′-H), 6.78 (d, J = 8.1 Hz, 1 H, 5′-H), 6.82 (dd, J = 8.2, 2.0 Hz, 1 H, 6′-H). – 13C NMR (125 MHz, CD3OD): δ = 78.5 (C-2), 66.1 (C-3), 27.1 (C-4), 156.0 (C-5), 94.9 (C-6), 156.6 (C-7), 94.5 (C-8), 156.3 (C-9), 98.7 (C-10), 130.9 (C-1′), 113.9 (C-2′), 144.5 (C-3′), 144.4 (C-4′), 114.5 (C-5′), 117.9 (C-6′).

3.3.4 Methyl citrate (4)

1H NMR (500 MHz, CD3OD, 25 °C, TMS): δ = 2.78 (d, J = 15.7 Hz, 2 H, 2-H), 2.93 (d, J = 15.7 Hz, 2 H, 4-H), 3.68 (s, 3 H, OMe). – 13C NMR (125 MHz, CD3OD): δ = 174.1 (C-1′), 171.9 (C-1 and C-5), 42.7 (C-2 and C-4), 73.2 (C-3), 51.6 (OMe).

3.3.5 Tovopyrifolin C (7)

1H NMR (500 MHz, C2D6SO, 25 °C, TMS) δ = 6.52 (dd, J = 7.6, 1.9 Hz, 1 H, 4-H), 7.34 (t, J = 7.7 Hz, 1 H, 6-H), 7.28 (dd, J = 7.6, 1.9 Hz, 1 H, 7-H), 7.64 (s, 1 H, 8-H), 3.86 (s, 3 H, OMe). – 13C NMR (125 MHz, CD3OD): δ = 155.3 (C-1), 131.5 (C-2), 159.2 (C-3), 94.7 (C-4), 153.8 (C-4a), 146.5 (C-5), 153.1 (C-6), 124.8 (C-7), 116.1 (C-8), 121.7 (C-8a), 182.1 (C-9), 104.0 (C-9a), 146.1 (C-10a), 60.7 (OMe).

3.3.6 Cowagarcinone B (8)

1H NMR (500 MHz, C3D6O, 25 °C, TMS): δ = 6.62 (s, 1 H, 4-H), 6.98 (s, 1 H, 5-H), 7.58 (s, 1 H, 8-H), 13.43 (s, 1 H, 1-OH), 4.01 (s, 6H, 7-OMe; 3-OMe), 6.32 (s, 1 H, 6-OH), 3.36 (d, J = 7.3 Hz, 2 H, 1′-H), 5.23 (brt, J = 7.3 Hz, 1 H, 2′-H), 1.80 (s, 3 H, 4′-H), 1.66 (s, 3-H, 5′-H). – 13C NMR (125 MHz, CD3OD): δ = 159.3 (C-1), 111.7 (C-2), 163.8 (C-3), 89.6 (C-4), 156.2 (C-4a), 102.5 (C-5), 152.4 (C-6), 144.3 (C-7), 104.6 (C-8), 113.6 (C-8a), 179.9 (C-9), 103.4 (C-9a), 152.5 (C-10a), 56.5(OMe-7), 55.9 (OMe-3), 21.4 (C-1′),122.1 (C-2′), 131.9 (C-3′), 17.8 (C-4′), 25.8 (C-5′).

3.3.7 α-Mangostin (9)

1H NMR (500 MHz, CD3OD, 25 °C, TMS): δ = 6.26 (s, 1 H, 4-H), 6.72 (s, 1 H, 5-H), 3.78 (s, 3 H, 7-OMe), 3.31 (d, J = 7.1 Hz, 2 H, 1′-H), 5.24 (m, 1 H, 2′-H), 1.68 (s, 3 H, 4′-H), 1.69 (s, 3 H, 5′-H), 4.11 (s, 2 H, 1″-H), 5.24 (m, 1 H, 2″-H), 1.80 (s, 3 H, 4″-H), 1.84 (s, 3 H, 5″-H). – 13C NMR (125 MHz, CD3OD): δ = 160.1 (C-1), 109.8 (C-2), 162.4 (C-3), 91.5 (C-4), 155.8 (C-4a), 101.2 (C-5), 154.8 (C-6), 143.5 (C-7), 137.3 (C-8), 111.1 (C-8a), 181.6 (C-9), 102.8 (C-9a), 156.5 (C-10a), 20.7 (C-1′), 123.9 (C-2′), 130.3 (C-3′), 24.3 (C-4′), 16.9 (C-5′), 26.0 (C-1″), 122.4 (C-2″), 130.4 (C-3″), 24.3 (C-4″), 16.4 (C-5″), 59.8 (OMe).

3.3.8 Lupeol (10)

1H NMR (500 MHz, CDCl3, 25 °C, TMS): δ = 3.21 (dd, J = 11.4, 4.9 Hz, 1 H, 3-H), 0.99 (s, 3 H, 23-H), 0.81 (s, 3 H, 24-H), 0.85 (s, 3 H, 25-H), 1.1 (s, 3 H, 26-H), 0.97 (s, 3 H, 27-H), 0.79 (s, 3 H, 28-H), 4.59 (d, J = 2.7 Hz, 1 H, 29a-H), 4.71 (d, J = 2.4 Hz, 1 H, 29b-H), 1.71 (s, 3 H, 30-H). – 13C NMR (125 MHz, CDCl3): δ = 38.7 (C-1), 27.5 (C-2), 79.0 (C-3), 38.9 (C-4), 55.3 (C-5), 18.3 (C-6), 34.3 (C-7), 40.8 (C-8), 50.4 (C-9), 37.2 (C-10), 20.9 (C-11), 25.1 (C-12), 38.1 (C-13), 42.8 (C-14), 27.4 (C-15), 35.6 (C-16), 43.0 (C-17), 48.3 (C-18), 48.0 (C-19), 151.0 (C-20), 29.9 (C-21), 40.0 (C-22), 28.0 (C-23), 15.4 (C-24), 16.1 (C-25), 16.0 (C-26), 14.6 (C-27), 18.0 (C-28), 109.3 (C-29), 19.3 (C-30).

3.3.9 Betulin (11)

1H NMR (500 MHz, C3D6O, 25 °C, TMS): δ = 3.15 (dd, J = 11.0, 5.3 Hz, 1 H, 3-H), 0.99 (s, 3 H, 23-H), 0.78 (s, 3 H, 24-H), 0.89 (s, 3 H, 25-H), 1.09 (s, 3 H, 26-H), 1.04 (s, 3 H, 27-H), 3.32 (d, J = 10.6 Hz, 1 H, 28a-H), 3.76 (d, J = 10.3 Hz, 1 H, 28b-H), 4.59 (brs, 1 H, 29a-H), 4.71 (brs, 1 H, 29b-H), 1.72 (s, 3 H, 30-H). – 13C NMR (125 MHz, C3D6O): δ = 38.7 (C-1), 27.4 (C-2), 77.7 (C-3), 38.7 (C-4), 55.5 (C-5), 18.2 (C-6), 34.3 (C-7), 40.9 (C-8), 50.5 (C-9), 37.1 (C-10), 20.7 (C-11), 25.4 (C-12), 37.3 (C-13), 42.6 (C-14), 27.4 (C-15), 29.4 (C-16), 47.8 (C-17), 48.7 (C-18), 47.9 (C-19), 150.8 (C-20), 29.8 (C-21), 34.0 (C-22), 27.1 (C-23), 15.1 (C-24), 15.7 (C-25), 15.6 (C-26), 14.3 (C-27), 59.0 (C-28), 109.0 (C-29), 18.4 (C-30).

3.3.10 Tovophylin A (12)

1H NMR (500 MHz, CDCl3, 25 °C, TMS): δ = 6.37 (s, 1 H, 4-H), 3.49 (d, J = 7.2 Hz, 2 H, 1′-H), 5.31 (dt, J = 13.3, 7.2 Hz, 1 H, 2′-H), 1.71 (s, 3 H, 4′-H), 1.90 (s, 3 H, 5′-H), 3.60 (d, J = 7.3 Hz, 2 H, 1″-H), 5.31 (dt, J = 13.3, 7.2 Hz, 2 H, 2″-H), 1.80 (s, 3 H, 4″-H), 1.87 (s, 3 H, 5″-H), 5.79 (d, J = 10.2 Hz, 1 H, 1‴-H), 8.03 (d, J = 10.2 Hz, 1 H, 2‴-H), 1.51 (s, 6 H, 4‴-H and 5‴-H), 13.79 (s, 1 H, 1-OH), 6.31 (s, 1 H, 6-OH), 6.12 (s, 1 H, 3-OH). – 13C NMR (125 MHz, CDCl3): δ = 160.4 (C-1), 108.2 (C-2), 161.7 (C-3), 93.4 (C-4), 155.3 (C-4a), 115.8 (C-5), 148.6 (C-6), 136.5 (C-7), 117.2 (C-8), 108.4 (C-8a), 182.9 (C-9), 103.7 (C-9a), 151.0 (C-10a), 22.6 (C-1′), 121.5 (C-2′), 131.3 (C-3′), 25.9 (C-4′), 18.0 (C-5′), 21.5 (C-1″), 121.1 (C-2″), 132.6 (C-3″), 25.8 (C-4″), 17.9 (C-5″), 136.0 (C-1‴), 121.0 (C-2‴), 76.4 (C-3‴), 27.4 (C-4‴ and C-5‴).

3.3.11 1,3,7-Trihydroxyxanthone (13)

1H NMR (500 MHz, CD3OD, 25 °C, TMS): δ = 6.19 (d, J = 2.1 Hz, 1 H, 2-H), 6.33 (d, J = 2.1 Hz, 1 H, 4-H), 7.38 (d, J = 9.0 Hz, 1 H, 5-H), 7.26 (dd, J = 9.0, 3.0 Hz, 1 H, 6-H), 7.50 (d, J = 2.9 Hz, 1 H, 8-H). – 13C NMR (125 MHz, CD3OD): δ = 165.8 (C-1), 97.5 (C-2), 163.3 (C-3), 93.4 (C-4), 158.4 (C-4a), 118.4 (C-5), 123.9 (C-6), 153.9 (C-7), 108.0 (C-8), 120.8 (C-8a), 180.4 (C-9), 102.3 (C-9a), 149.8 (C-10a).

3.3.12 9-Hydroxycalabaxanthone (14)

1H NMR (500 MHz, CDCl3, 25 °C, TMS): δ = 6.17 (s, 1 H, 4-H), 6.76 (s, 1 H, 5-H), 6.66 (d, J = 10.0 Hz, 1 H, 1′-H), 5.49 (d, J = 10.0 Hz, 1 H, 2′-H), 1.39 (s, 6 H, 4′-H and 5′-H), 4.02 (s, 2 H, 1″-H), 5.19 (brt, J = 6.3 Hz, 1 H, 2″-H), 1.62 (s, 3 H, 4″-H), 1.76 (s, 3 H, 5″-H), 13.62 (s, 1 H, 1-OH), 3.73 (s, 3 H, 7-OMe). – 13C NMR (125 MHz, CDCl3): δ = 157.9 (C-1), 104.5 (C-2), 159.9 (C-3), 94.2 (C-4), 156.3 (C-4a), 101.7 (C-5), 154.6 (C-6), 142.7 (C-7), 137.0 (C-8), 112.2 (C-8a), 182.0 (C-9), 103.7 (C-9a), 155.8 (C-10a), 115.7 (C-1′), 127.2 (C-2′), 77.9 (C-3′), 28.3 (C-4′ and C-5′), 26.6 (C-1″), 123.1 (C-2″), 132.2(C-3″), 25.8 (C-4″), 18.2 (C-5″), 62.1 (OMe).

3.4 In vitro antileishmanial activity

L. donovani 1S(MHOM/SD/62/1S) promastigotes were cultivated at 28 °C in axenic M199 culture medium (Sigma Aldrich) supplemented with 10% heat-inactivated fetal bovine serum (FBS) (Sigma Aldrich) and 1% streptomycin/penicillin (Sigma Aldrich). The antileishmananial activity of test samples were determined as previously described [27], using the resazurin-based assay. Compounds were serially diluted in incomplete M199 medium and 10 μL of each compound were introduced in 90 μL of L. donovani promastigotes (4 × 105 parasites) from an exponential phase culture in complete medium. They were all screened at final concentrations of 100–0.16 μg mL−1 for extracts and fractions and 5–0.08 μg mL−1 for compounds and test plates were incubated for 28 h at 28 °C, followed by the addition of 1 mg mL−1 resazurin. The negative and positive controls were 0.1% DMSO and amphotericin B (10–0.016 μg mL−1), respectively. After an additional incubation of 44 h, plates were then read on a Magelan Infinite M200 fluorescence multi-well plate reader (Tecan) at an excitation and an emission wavelength of 530 and 590 nm, respectively. For each sample, growth percentages were calculated and dose-response curves were constructed to determine the 50% inhibitory concentration (IC50) using the GraphPad (version 5.0) software. The results showed in Table 1 and 2, were discussed according to the classification established by Camacho et al. (2003) (IC50 < 10 μg mL−1, extract is highly active; 10 < IC50 < 50 μg mL−1, extract is good active; 50 < IC50 < 100 μg mL−1, extract is moderately active; IC50 > 100 μg mL−1, extract is inactive) [28].

3.5 In vitro antibacterial assay

Antibacterial activity assays were conducted on a total of six bacterial strains, two from American type culture collection, E. coli ATCC 25322 and S. pneumoniae ATCC 491619, one from BEI resources namely P. aeruginosa HM801 and finally three clinical isolate strains from laboratory collection namely S. typhi (CPC and CHU), E. cloacae (CPC), and S. aureus (CPC). They were assessed for their susceptibility to extracts/compounds. The tests were performed in duplicate, following the method by Eloff (1998) [29]. In a 96-well microplate, 100 μL of sterile culture broth (MHB) were introduced. Then 100 μL of each stock sample solution (2000 μg mL−1) were added to the first wells and then distributed to all other wells, with concentrations ranging from 3.8 to 500 μg mL−1 and < 0.15–500 μg mL−1 for ciprofloxacin. Then 100 μL of liquid culture medium (MHB) inoculated with the test organism (2 × 106 CFU mL−1) were introduced into the wells in order to obtain a final concentration of 106 CFU mL−1. Ciprofloxacin was used as reference. The negative controls consisted of wells containing only the culture medium, and wells containing a mixture of culture broth and test organism. The culture microplates were covered and incubated at 37 °C (18 h). Twenty microliters (20 μL) of resazurin were then introduced into both and then incubated again at 37 °C for 24 h [30]. The antibacterial activity of compound is strong, moderate and weak if the MIC of the plants is ≤ 10 μg mL−1, 10 < MIC ≤ 100 μg mL−1 and > 100 μg mL−1, respectively. Whereas, the activity of plant extracts is classified as significant if (MIC < 100 μg mL−1), moderate if (100 < MIC ≤ 625 μg mL−1) and weak if (MIC > 625 μg mL−1) [31].

4 Supporting information

The isolation protocols, 1H and 13C NMR spectra are given as supplementary material available online (https://doi.org/10.1515/znb-2021-0077).

Corresponding author: Jean Jules Kezetas Bankeu, Department of Chemistry, Faculty of Science, The University of Bamenda, P. O. Box 39, Bambili, Cameroon, E-mail: ; and Bruno Ndjakou Lenta, Department of Chemistry, Higher Teacher Training College, University of Yaoundé I, P. O. Box 47, Yaoundé, Cameroon, E-mail:

  1. Author contribution: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

  2. Research funding: This work was supported by the Yaoundé-Bielefeld Bilateral Graduate School of Natural Products with Antiparasite and Antibacterial activity (YaBiNaPA) project, financially supported by Deutscher Akademischer Austauschdienst (DAAD) [grant number 57316173].

  3. Conflict of interest statement: The authors declare no competing financial interest.

References

1. Boer, D. M., Argaw, D., Jannin, J., Alvar, J. Clin. Microbiol. Infect. 2011, 17, 1471–1477, https://doi.org/10.1111/j.1469-0691.2011.03635.x.Search in Google Scholar PubMed

2. Akhoundi, M., Kuhls, K., Cannet, A., Votýpka, J., Marty, P., Delaunay, S. D. P. PLoS Neglected Trop. Dis. 2016, 10, e0004349, https://doi.org/10.1371/journal.pntd.0004349.Search in Google Scholar PubMed PubMed Central

3. Torres-Guerrero, E., Quintanilla-Cedillo, M. R., Ruiz-Esmenjaud, J., Arenas, R. F1000 Research 2017, 6, 1–38, https://doi.org/10.12688/f1000research.11120.1.Search in Google Scholar PubMed PubMed Central

4. Singh, N., Mishra, B. B., Bajpai, S., Singh, R. K., Tiwari, V. K. Bioorg. Med. Chem. 2014, 22, 18–45, https://doi.org/10.1016/j.bmc.2013.11.048.Search in Google Scholar PubMed

5. Lenta, N. B., Vonthron-Sénécheau, C., Weniger, B., Devkota, K. P., Ngoupayo, J., Kaiser, M., Naz, Q., Choudhary, M. I., Tsamo, E., Sewald, N. Molecules 2007, 12, 1548–1557, https://doi.org/10.3390/12081548.Search in Google Scholar PubMed PubMed Central

6. Aissi, M. V., Tchobo, F. P., Natta, A. K., Piombo, G., Villeneuve, P., Sohounhloué, D. C. K., Soumanou, M. M. OCL: Oilseeds Fats, Crops Lipids 2011, 18, 384–392, https://doi.org/10.1051/ocl.2011.0423.Search in Google Scholar

7. Raponda-Walker, A., Sillans, R. Les Plantes Utiles du Gabon; Paul Lechevalier: Paris, 1961; pp. 614.10.3109/13880206109066644Search in Google Scholar

8. Iwu, M. M. Handbook of African Medicinal Plants; CRC Press: London, 1993; pp. 36–37.Search in Google Scholar

9. Ee, G. C. L., Lim, C. K., Ong, G. P., Sukari, M. A., Lee, H. L. J. Asian Nat. Prod. Res. 2006, 8, 567–570, https://doi.org/10.1080/10286020500172335.Search in Google Scholar PubMed

10. Ninh, T. S., Nguyen, T. T. H., Pham, V. C., Le, T. A., Nguyen, T. T., Ba, T. C. Vietnam J. Chem. 2020, 58, 343–348.10.1002/vjch.2019000183Search in Google Scholar

11. Usman, A., Thoss, V., Nur-e-Alam, M. Am. J. Org. Chem. 2016, 6, 81–85.Search in Google Scholar

12. Li, M., Wang, Y., Fu, D., Liu, X. Acta Crystallogr. 2007, E63, o4497, https://doi.org/10.1107/s1600536807052828.Search in Google Scholar

13. Khatun, M., Billah, M., Quader, M. A. Dhaka Univ. J. Sci. 2012, 60, 5–10, https://doi.org/10.3329/dujs.v60i1.10327.Search in Google Scholar

14. Nyigo, V. A., Peter, X., Mabiki, F., Malebo, H. M., Mdegela, R. H., Fouche, G. J. Phytopharm. 2016, 5, 100–104, https://doi.org/10.31254/phyto.2016.5302.Search in Google Scholar

15. Ee, G. C. L., Jong, V. Y. M., Sukari, M. A., Rahmani, M., Kua, A. S. M. J. Sci. Technol. 2009, 17, 307–312.Search in Google Scholar

16. Mahabusarakam, W., Chairek, P., Taylor, W. C. Phytochemistry 2005, 66, 1148–1153, https://doi.org/10.1016/j.phytochem.2005.02.025.Search in Google Scholar

17. Upegui, Y., Robledo, S. M., Gil Romero, J. F., Quiñones, W., Archbold, R., Torres, F., Escobar, G., Narino, B., Echeverri, F. Phytother Res. 2015, 298, 1195–1201, https://doi.org/10.1002/ptr.5362.Search in Google Scholar

18. Mahato, S. B., Kundu, P. A. Phytochemistry 1994, 37, 1517–1575, https://doi.org/10.1016/s0031-9422(00)89569-2.Search in Google Scholar

19. Goncalves De Oliveira, W., Gottlieb, R., Lins Mesquita, A. A. Phytochemistry 1972, 11, 3323–3325.https://doi.org/10.1016/s0031-9422(00)86400-6.Search in Google Scholar

20. Mukulesh, M., Vedavati, G. P., Narshinha, P. A. J. Org. Chem. 2006, 71, 4992–4995.10.1021/jo0606655Search in Google Scholar PubMed

21. Silva, E. M., Araújo, R. M., Freire-Filha, L. G., Silveira, E. R., Lopes, N. P., de Paula, J. E., Braz-Filhof, R., Espindola, L. S. J. Braz. Chem. Soc. 2013, 24, 1314–1321.Search in Google Scholar

22. Azebaze, A. G. B., Ouahouo, B. M. W., Vardamides, J. C., Valentin, A., Kuete, V., Acebey, L., Beng, V. P., Nkengfack, A. E., Meyer, M. Chem. Nat. Compd. 2008, 44, 582–587, https://doi.org/10.1007/s10600-008-9141-9.Search in Google Scholar

23. Al-Massarani, S., El Gamal, A., Al-Musayeib, N., Mothana, R., Basudan, O., Al-Rehaily, A., Farag, M., Assaf, M., Tahir, K., Maes, L. Molecules 2013, 18, 10599–10608, https://doi.org/10.3390/molecules180910599.Search in Google Scholar PubMed PubMed Central

24. Koh, J.-J., Qiu, S., Zou, H., Lakshminarayanan, R., Li, J., Zhou, X., Tang, C., Saraswathi, P., Verma, C., Tan, D. T. H., Tan, A. L., Liu, S., Beuerman, R. W. Biochim. Biophys. Acta 2013, 1828, 834–844, https://doi.org/10.1016/j.bbamem.2012.09.004.Search in Google Scholar PubMed

25. Araújo, J., Fernandes, C., Pinto, M., Tiritan, M. E. Molecules 2019, 24, 314, https://doi.org/10.3390/molecules24020314.Search in Google Scholar

26. Dharmaratne, H. R. W., Sakagami, Y., Piyasena, K. G. P., Thevanesam, V. Nat. Prod. Res. 2013, 27, 938–941, https://doi.org/10.1080/14786419.2012.678348.Search in Google Scholar

27. Siqueira-Neto, J. L., Song, O.-R., Oh, H., Sohn, J.-H., Yang, G., Nam, J., Jang, J., Cechetto, J., Lee, C. B., Moon, S., Genovesio, A., Chatelain, E., Christophe, T., Freitas-Junior, L. H. PLoS Neglected Trop. Dis. 2010, 4, e675, https://doi.org/10.1371/journal.pntd.0000675.Search in Google Scholar

28. Camacho, M. D. R., Phillipson, J. D., Croft, S. L., Solis, P. N., Marshall, S. J., Ghazanfar, S. A. J. Ethnopharmacol. 2003, 89, 185–191, https://doi.org/10.1016/s0378-8741(03)00269-1.Search in Google Scholar

29. Eloff, J. N. Planta Med. 1998, 64, 711–713, https://doi.org/10.1055/s-2006-957563.Search in Google Scholar PubMed

30. Mativandlela, S. P. N., Lall, N., Meyer, J. J. M. S. Afr. J. Bot. 2006, 72, 232–237, https://doi.org/10.1016/j.sajb.2005.08.002.Search in Google Scholar

31. Kuete, V., Alibert-Franco, S., Eyong, K. O., Ngameni, B., Folefoc, G. N., Nguemeving, J. R., Tangmouo, J. G., Fotso, G. W., Komguem, J., Ouahouo, B. M. W., Bolla, J. M., Chevalier, J., Ngadjui, B. T., Nkengfack, A. E. Int. J. Antimicrob. Agents 2011, 37, 156–161, https://doi.org/10.1016/j.ijantimicag.2010.10.020.Search in Google Scholar PubMed

Supplementary Material

The online version of this article offers supplementary material (https://doi.org/10.1515/znb-2021-0077).

Received: 2021-05-26
Accepted: 2021-08-31
Published Online: 2021-10-21
Published in Print: 2022-01-27

© 2021 Walter de Gruyter GmbH, Berlin/Boston

Articles in the same Issue

  1. Frontmatter
  2. In this issue
  3. Research Articles
  4. A new phenanthrene derivative from Entada abyssinica with antimicrobial and antioxidant properties
  5. Antileishmanial, antibacterial and cytotoxicity activity of the extracts, fractions, and compounds from the fruits and stem bark extracts of Pentadesma butyracea Sabine
  6. Two 2D Co(II)/Mn(II) coordination polymers based on the quinoline-2,3-dicarboxylate ligand: synthesis, crystal structure, and fluorescence properties
  7. Synthesis, hydrogen bond interactions and crystal structure elucidation of some stable 2H-imidazolium salts
  8. Molecular and crystal structure of a copper(II) complex of sildenafil
  9. A convenient approach for the electrochemical bromination and iodination of pyrazoles
  10. Furanone-functionalized benzothiazole derivatives: synthesis, in vitro cytotoxicity, ADME, and molecular docking studies
  11. Si⋯O proximity in imidosilanes – absence of orbital interactions
  12. A new luminescent metal-organic framework with 2,6-di(1H-imidazol-1-yl)naphthalene and biphenyl-3,4′,5-tricarboxylic acid
  13. Chalcogenative spirocyclization of N-aryl propiolamides with diselenides/disulfides promoted by Selectfluor
  14. [Msim]CuCl3: as an efficient catalyst for the preparation of 5-amino-1H-pyrazole-4-carbonitriles by anomeric based oxidation
  15. Synthesis and structure of an asymmetrical sila[1]magnesocenophane
https://doi.org/10.1515/znb-2021-0077
Keywords for this article
antibacterial; antileishmanial; clusiaceae; cytotoxicity; Pentadesma butyracea

Articles in the same Issue

  1. Frontmatter
  2. In this issue
  3. Research Articles
  4. A new phenanthrene derivative from Entada abyssinica with antimicrobial and antioxidant properties
  5. Antileishmanial, antibacterial and cytotoxicity activity of the extracts, fractions, and compounds from the fruits and stem bark extracts of Pentadesma butyracea Sabine
  6. Two 2D Co(II)/Mn(II) coordination polymers based on the quinoline-2,3-dicarboxylate ligand: synthesis, crystal structure, and fluorescence properties
  7. Synthesis, hydrogen bond interactions and crystal structure elucidation of some stable 2H-imidazolium salts
  8. Molecular and crystal structure of a copper(II) complex of sildenafil
  9. A convenient approach for the electrochemical bromination and iodination of pyrazoles
  10. Furanone-functionalized benzothiazole derivatives: synthesis, in vitro cytotoxicity, ADME, and molecular docking studies
  11. Si⋯O proximity in imidosilanes – absence of orbital interactions
  12. A new luminescent metal-organic framework with 2,6-di(1H-imidazol-1-yl)naphthalene and biphenyl-3,4′,5-tricarboxylic acid
  13. Chalcogenative spirocyclization of N-aryl propiolamides with diselenides/disulfides promoted by Selectfluor
  14. [Msim]CuCl3: as an efficient catalyst for the preparation of 5-amino-1H-pyrazole-4-carbonitriles by anomeric based oxidation
  15. Synthesis and structure of an asymmetrical sila[1]magnesocenophane
Sign up now to receive a 20% welcome discount
Institutional Access
How does access work?
Have an idea on how to improve our website?
Please write us.
© 2025 De Gruyter Brill
Downloaded on 24.9.2025 from https://www.degruyterbrill.com/document/doi/10.1515/znb-2021-0077/html?lang=en
Scroll to top button