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Communication

New Derivatives of Lupeol and Their Biological Activity

1
Department of Organic Chemistry, Faculty of Chemistry, Ho Chi Minh City University of Education, Ho Chi Minh City 72711, Vietnam
2
Faculty of Environmental Science, Sai Gon University, Ho Chi Minh City 72711, Vietnam
3
Faculty of Basic Sciences, University of Medicine and Pharmacy at Ho Chi Minh City, 217 Hong Bang Street, Dist. 5, Ho Chi Minh City 700000, Vietnam
4
CirTech Institute, HUTECT University, Ho Chi Minh City 72324, Vietnam
5
Research Unit in Natural Products Chemistry and Bioactivities, Faculty of Science and Technology, Thammasat University Lampang Campus, Lampang 52190, Thailand
*
Authors to whom correspondence should be addressed.
Molbank 2021, 2021(4), M1306; https://doi.org/10.3390/M1306
Received: 16 November 2021 / Revised: 26 November 2021 / Accepted: 6 December 2021 / Published: 10 December 2021
(This article belongs to the Special Issue Synthesis of Flavonoids or Other Nature-Inspired Small Molecules)

Abstract

:
The natural product lupeol (1) was isolated from Bombax ceiba leaves, which were used as starting material in the semisynthetic approach. Three new derivatives (2a, 2b, and 3) were synthesized using oxidation and aldolization. Their chemical structures were elucidated by spectroscopic analyses (HRESIMS and NMR). Compounds 3 showed significant α-glucosidase inhibition with an IC50 value of 202 µM, whereas 2a and 2b were inactive.

1. Introduction

Diabetes mellitus (DM) causes high blood glucose after the consumption of a carbohydrate-enriched diet, leading to hyperglycemia. Uncontrolled diabetes is manifested by a very high rise in triglycerides and fatty acid levels [1]. Diverse antidiabetic drugs derived from synthetic compounds are of interest to chemists. However, these synthetic drugs come with several serious complications [1]. Due to the limitations associated with the use of existing synthetic antidiabetic drugs, the search for newer antidiabetic agents from natural sources continues. Lupeol is a pharmacologically active pentacyclic triterpenoid found in several medicinal plants worldwide [2]. It has several potential medicinal properties and is found in a variety of botanical sources [3]. Notably, lupeol has been reported to selectively target diseased and unhealthy human cells, while sparing normal and healthy cells [4]. Dozens of novel lupeol derivatives were synthesized and screened for their in vivo antihyperglycemic activity [5,6]. Most derivatives lowered the blood glucose levels, in a sucrose-challenged streptozotocin-induced diabetic rat (STZ-S) model [5]. To continue our ongoing search for highly efficient antidiabetic agents from derivatized lupeol [6,7], we herein describe the synthesis of lupeol derivatives 2, 2a, 2b, and 3 (Figure 1). The structures of all the obtained compounds were characterized by 1H, 13C NMR, and HRESIMS. All derivatives were evaluated for α-glucosidase inhibition.

2. Results and Discussion

2.1. Synthesis

Lupeol was isolated from the Vietnamese plant Bombax ceiba, following our previously reported procedure [8]. Lupeol was transformed to products 2, 2a, and 2b using oxidation with Oxone®, a potassium triple-salt (KHSO5·1/2KHSO4·1/2K2SO4) [6,9]. The conditions followed our previously reported method [6], with slight modifications. Both 2a and 2b had the same molecular formula as C32H52O4. Comparison of NMR data of 2a/2b and 1 indicated that oxidation occurred. The 1H NMR spectrum of 2a/2b showed differences with 1: the downfield methine at δH 8.11, two oxymethines at δH 5.26 and 4.48, and a doublet methyl at δH 1.22. These signals indicated that the isopropenyl group of 1 was transformed to a 2-formylethyl group at C-19. Moreover, the downfield signal of H-3 (δH 4.48) indicated that 3-OH was esterified by acetic acid. The 13C NMR spectrum of 2a/2b showed one carbonyl ester at δC 171.1, one formyl group at δC 163.7 and two oxygenated carbons at δC 81.1 and 72.7, supporting the previous findings. Interestingly, 2a and 2b are C-20 epimers. Corbett and co-workers [10,11] indicated the method to define the absolute configuration of C-20 of lupane-type triterpenes. Particularly, the (20S) and (20R) isomers exhibited differences in the chemical shifts of C-19, C-20, C-29, and C-30, especially C-30. According to Corbett et al., 2a, having C-30 at δC 20.1, would have a 20R configuration. On the other hand, 2b would have the 20S configuration due to the lower chemical shift of C-30 at δC 14.2.
Compound 2 was further aldolized with 4-bromobenzaldehyde to afford compound 3. Compound 3 had the same molecular formula as C36H51BrO2, determined by a protonated ion peak at m/z 595.3188 in HRESIMS. Comparison of 1D NMR data of 2 and 3 indicated obvious differences. The first difference is the presence of a 1,4-disubstituted benzenoid characterized by two ortho-coupled protons at δH 7.51 and 7.42, and a trans double bond at δH 6.75 and 7.46. This was confirmed by the disappearance of a methyl ketone group at δH 2.15 (CH3-29). This finding indicated that the aldolization occurred exclusively at C-29. The second difference was in the 13C NMR spectrum. This spectrum showed the presence of seven aromatic carbons at δC 141.0 (C-1), 133.9 (C-5′), 132.3 (C-2′), 129.8 (C-3′,7′), and 126.9 (C-4′, 6′), supporting the reaction at C-29.

2.2. α-Glucosidase Inhibition of 2a, 2b, and 3

Compounds 2a, 2b, and 3 were evaluated for α-glucosidase inhibition. Only compound 3 exhibited moderate α-glucosidase inhibition with an IC50 value of 202 µM, compared with an acarbose-positive control (IC50 360 μM). Other compounds were inactive.

3. Materials and Methods

3.1. Materials

Reagents and solvents were obtained from commercial suppliers and were used without further purification. Column chromatography was carried out using Merck Kieselgel 60 silica gel (particle size: 32–63 Å). Analytical TLC was performed using Merck precoated silica gel 60 F-254 sheets.
NMR spectroscopic data were acquired on Bruker Avance III apparatus at 500 MHz for 1H NMR and 125 MHz for 13C NMR. HRESIMS spectra were recorded on a Bruker MICROTOF-Q 10187.
Extraction and Isolation. The air-dried Bombax ceiba leaves (4 kg) were ground into powder and exhaustively extracted at room temperature with MeOH (2 × 10 L). The filtered solution was evaporated under reduced pressure to afford a residue (473.4 g). This crude extract was subsequently partitioned using solvents of n-hexane and EtOAc to yield n-hexane (40 g) and EtOAc (88 g) extracts. The n-hexane extract was fractionated by silica gel column chromatography (CC), eluted with n-hexane–EtOAc (isocratic, 10:1, v/v), to produce five fractions (H1-H5). Fraction H2 (15 g) was rechromatographed by silica gel CC using n-hexane–CHCl3 (isocratic, 12:1, v/v) as eluent to afford lupeol (1) (1.5 g).

3.2. Synthesis Procedure

Synthesis of 2, 2a, and 2b: Lupeol (1, 200 mg, 0.469 mmol) was oxidized with Oxone® (951 mg, 1.548 mmol) in acetic acid (40 mL) at 100 °C for 3 h. The mixture was stirred and continuously monitored by TLC. The mixture was extracted with EtOAc–water (1:1) to gain the organic layer. This solution was evaporated to afford a residue. Then, the residue was purified by silica gel CC to give compounds 2, 2a, and 2b.
Compound 2. Isolated yield: 74.6 mg (37%), white solid. 1H and 13C NMR data were consistent with those reported previously [6].
(3aR,5aR,5bR,7aR,9S,11aR,11bR,13aR,13bS)-1-((S)-1-(formyloxy)ethyl)-3a,5a,5b,8,8,11a-hexamethylicosahydro-1H-cyclopenta[a]chrysen-9-yl acetate (2a). Isolated yield: 9.4 mg (4%), white solid. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.05 (3H, s, CH3-2′), 8.00 (1H, s, OCHO-29), 5.33 (1H, m, H-20), 4.48 (1H, dd, J = 10.5, 6.0 Hz, H-3), 2.13 (1H, m, H-3), 1.18 (3H, d, J = 6.5 Hz, CH3-30), 1.03 (3H, s, CH3-26), 0.90 (3H, s, CH3-27), 0.87 (3H, s, CH3-25), 0.85 (3H, s, CH3-23), 0.84 (3H, s, CH3-24), 0.79 (1H, d, J = 9.5 Hz, H-5), 0.76 (3H, s, CH3-28). 13C NMR (125 MHz, CDCl3, δ, ppm): 171.2 (C-6′), 21.5 (C2′), 161.6 (C-29), 81.1 (C-3), 73.4 (C-20), 55.5 (C-5), 50.2 (C-9), 48.8 (C-18), 43.5 (C-17), 43.0 (C-14), 42.6 (C-19), 41.0 (C-8), 40.5 (C-22), 38.5 (C-4), 38.0 (C-1), 37.3 (C-13), 37.2 (C-10), 35.5 (C-16), 34.4 (C-7), 29.9 (C-21), 28.1 (C-23), 27.3 (C-15), 27.1 (C-2), 23.8 (C-12), 21.0 (C-11), 18.4 (C-6), 18.1 (C-28), 16.7 (C-25), 16.4 (C-26), 16.1 (C-24), 14.4 (C-27), 14.2 (C-30). HRESIMS calcd C32H52NaO4 ([M+Na]+): 523.3732, found: 523.3763.
(3aR,5aR,5bR,7aR,9S,11aR,11bR,13aR,13bS)-1-((R)-1-(formyloxy)ethyl)-3a,5a,5b,8,8,11a-hexamethylicosahydro-1H-cyclopenta[a]chrysen-9-yl acetate (2b). Yield: 9.4 mg (5%), white solid. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.04 (3H, s, H-2′), 8.11 (1H, s, OCHO-29), 5.26 (1H, m, H-20), 4.48 (1H, dd, J = 11.5, 5.5 Hz, H-3), 2.31 (1H, m, H-19), 1.22 (3H, d, J = 6.5 Hz, CH3-30), 1.03 (3H, s, CH3-26), 0.86 (3H, s, CH3-25), 0.85 (3H, s, CH3-27), 0.85 (3H, s, CH3-23), 0.84 (3H, s, CH3-24), 0.77 (1H, d, J = 2.0 Hz, H-5), 0.75 (3H, s, CH3-28). 13C NMR (125 MHz, CDCl3, δ, ppm): 171.1 (C-6′), 21.5 (C2′), 163.7 (C-29), 81.1 (C-3), 72.7 (C-20), 55.5 (C-5), 50.0 (C-9), 47.1 (C-18), 44.4 (C-19), 43.2 (C-17), 43.0 (C-14), 41.0 (C-8), 40.1 (C-22), 38.5 (C-4), 38.0 (C-1), 37.5 (C-13), 37.3 (C-10), 35.3 (C-16), 34.4 (C-7), 29.9 (C-21), 28.1 (C-23), 27.4 (C-15), 26.9 (C-2), 23.9 (C-12), 21.0 (C-11), 20.1 (C-30), 18.4 (C-6), 18.1 (C-28), 16.7 (C-25), 16.3 (C-26), 16.1 (C-24), 14.4 (C-27). HRESIMS calcd C32H52NaO4 ([M+Na+H2O]+): 541.3870, found: 541.3869.
Synthesis of 3: Compound 2 (70 mg, 0.163 mmol) together with NaOH (35 mg, 0.875 mmol) in ethanol (7 mL) was stirred at 55 °C for 15 min. Then, 4–bromobenzaldehyde (64.35 mg, 0.35 mmol) was added to the mixture. The reaction was performed at 55 °C for 2 h. The mixture was extracted with EtOAc–water (1:1, v/v) to gain the organic layer. This solution was applied to silica gel CC using the gradient system of n-hexane–EtOAc (10:1, v/v) to obtain compound 3. Isolated yield: 68 mg (48%), white solid.
(E)-3-(4-bromophenyl)-1-((1R,3aR,5aR,5bR,9S,11aR)-9-hydroxy-3a,5a,5b,8,8,11a-hexamethylicosahydro-1H-cyclopenta[a]chrysen-1-yl)prop-2-en-1-one (3): 1H NMR (500 MHz, CDCl3, δ, ppm): 7.51 (2H, d, J = 8.5 Hz, H-3′,7′), 7.46 (1H, d, J = 16.0 Hz, H-6′), 7.42 (2H, d, J = 8.5 Hz, H-4′,6′), 6.75 (1H, d, J = 16.0 Hz, H-29), 3.19 (1H, dd, J = 11.2, 4.8 Hz, H-3), 2.87 (1H, td, J = 11.5, 6.0 Hz, H-19), 1.02 (3H, s, CH3-26), 0.98 (3H, s, CH3-27), 0.96 (3H, s, CH3-23), 0.84 (3H, s, CH3-24), 0.80 (3H, s, CH3-25), 0.75 (3H, s, CH3-28). 13C NMR (125 MHz, CDCl3, δ, ppm): 204.1 (C-20), 141.0 (C-6′), 133.9 (C-5′), 132.3 (C-3′,7′), 129.8 (C-4′,6 ‘), 126.9 (C-29), 124.7 (C-2′), 79.0 (C-3), 55.4 (C-5), 50.4 (C-9), 50.1 (C-18), 43.3 (C-17), 42.9 (C-14), 40.9 (C-8), 40.3 (C-22), 39.0 (C-4), 38.8 (C-1), 37.3 (C-10), 35.2 (C-16), 34.3 (C-7), 28.7 (C-15), 28.1 (C-23), 27.9 (C-2), 27.5 (C-12), 21.1 (C-11), 18.4 (C-6), 18.3 (C-28), 16.2 (C-26), 16.0 (C-25), 15.5 (C-24), 14.6 (C-27). HRESIMS calcd C36H52BrO2 ([M−H]): 595.3151, found: 595.3188.

3.3. α-Glucosidase Inhibitory Assay

The α-glucosidase (0.2 U/mL) and substrate (5.0 mM p-nitrophenyl-α-D-glucopyranoside) were dissolved in 100 mM pH 6.9 sodium phosphate buffer [12]. The inhibitor (50 µL) was preincubated with α-glucosidase; then, the substrate (40 µL) was added to the reaction mixture. The enzymatic reaction was carried out at 37 °C for 20 min and stopped by the addition of 0.2 M Na2CO3 (130 μL). Enzymatic activity was quantified by measuring absorbance at 405 nm. All samples were analyzed in triplicate at five different concentrations around the IC50 values, and the mean values were retained. The inhibition percentage (%) was calculated as follows: Inhibition (%) = [1 − (Asample/Acontrol)] × 100.

4. Conclusions

Three new derivatives, 2a, 2b, and 3, from the natural product lupeol have been synthesized via oxidation and aldolization routes and evaluated for their α-glucosidase inhibition. Synthetic compound 3 showed much stronger α-glucosidase inhibitory activity (IC50 202 μM) than acarbose (IC50 360 μM). Synthetic products 2a and 2b, which lacked the 3-OH group, exhibited lower activity than 3 toward α-glucosidase. This result confirmed that this substituted group might be involved in α-glucosidase inhibition.

Supplementary Materials

The following are available online. Copies of HRESIMS and NMR spectra for compound 2a, 2b, and 3.

Author Contributions

Conceptualization, T.-H.D. and J.S. methodology, T.-H.D. and J.S.; formal analysis, H.-T.-T.L., T.-H.D. and J.S.; investigation, T.-H.-T.N., H.-T.-T.L., Q.-C.C., T.-H.D. and Q.-T.-P.T.; data curation, N.-K.-T.P. and N.-H.N.; writing—original draft preparation, T.-H.D. and J.S.; writing—review and editing, T.-H.D. and J.S.; supervision, T.-H.D.; project administration, T.-H.D. All authors have read and agreed to the published version of the manuscript.

Funding

The study was funded by The Youth Incubator for Science and Technology Programme, managed by the Youth Development Science and Technology Center—Ho Chi Minh Communist Youth Union and Department of Science and Technology of Ho Chi Minh City (38/2020/HĐ-KHCNT-VƯ). This work was also supported by Thammasat University Research Unit in Natural Products Chemistry and Bioactivities.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

The data for the compounds presented in this study are available in the Supplementary Materials of this paper.

Conflicts of Interest

The authors declare no conflict of interest.

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Figure 1. Synthesis of 2, 2a, 2b, and 3 from lupeol (1).
Figure 1. Synthesis of 2, 2a, 2b, and 3 from lupeol (1).
Molbank 2021 m1306 g001
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Le, H.-T.-T.; Chau, Q.-C.; Duong, T.-H.; Tran, Q.-T.-P.; Pham, N.-K.-T.; Nguyen, T.-H.-T.; Nguyen, N.-H.; Sichaem, J. New Derivatives of Lupeol and Their Biological Activity. Molbank 2021, 2021, M1306. https://doi.org/10.3390/M1306

AMA Style

Le H-T-T, Chau Q-C, Duong T-H, Tran Q-T-P, Pham N-K-T, Nguyen T-H-T, Nguyen N-H, Sichaem J. New Derivatives of Lupeol and Their Biological Activity. Molbank. 2021; 2021(4):M1306. https://doi.org/10.3390/M1306

Chicago/Turabian Style

Le, Hoang-Thuy-Tien, Quoc-Cuong Chau, Thuc-Huy Duong, Quyen-Thien-Phuc Tran, Nguyen-Kim-Tuyen Pham, Thi-Hoai-Thu Nguyen, Ngoc-Hong Nguyen, and Jirapast Sichaem. 2021. "New Derivatives of Lupeol and Their Biological Activity" Molbank 2021, no. 4: M1306. https://doi.org/10.3390/M1306

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