Two New Sesquiterpene Glycosides from Fissistigma balansae Merr

Abstract

Objective

Plants of Fissistigma balansae have been used for the treatment of stomachache in Vietnamese traditional medicinal remedies. This study reports 2 new sesquiterpene glycosides from the methanol extract of its branches.

Methods

The sample was extracted with MeOH and the crude extract obtained was suspended in H₂O and extracted with EtOAc. This extract was fractionated by various chromatographic methods to yield 2 new sesquiterpene glycosides. The structures of the isolates were determined by NMR spectroscopy and HRESI mass spectra.

Results

From the methanol extract of F balansae branches, 2 new sesquiterpene glycosides, named fissbalanoside A (1) and fissbalanoside B (2), have been isolated and their chemical structures evaluated. Compounds 1 and 2 weakly inhibited NO production with IC₅₀ values of 52.12  ±  1.78 and 65.27  ±  1.82 µM, respectively.

Introduction

The genus Fissistigma Griff., family Annonaceae, comprised about 76 species worldwide, which are woody climbers distributed in Asian countries. These plants are used in folk medicine to cure arthritis and stomachache pain. ¹ Fissistigma balansae Merr. is a climbing shrub, growing in Son Tay, Ba Vi, Phu Tho, Vinh Phu, Ha Tay, and Lam Dong provinces of Vietnam.^2,3 In previous papers, protoberberine alkaloids, ⁴ aporphine alkaloids, ⁵ dioxoaporphines and aristolactams, ⁶ and flavonoids and flavonoid glucosides ⁷ have been reported from this plant. This paper reports 2 previously undescribed sesquiterpene glycosides from the methanol extract of F balansae branches and their NO production effects.

Results and Discussion

Compound 1 (Figure 1) has a molecular formula of C₂₁H₃₆O₇ as determined from its HR-ESI-MS (m/z 435.2146 [M  +  ³⁵Cl]⁻, calcd for [C₂₁H₃₆O₇³⁵Cl]⁻: 435.2155 and m/z 437.2131 [M  +  ³⁷Cl]⁻, calcd for [C₂₁H₃₆O₇³⁷Cl]⁻: 437.2125). The ¹³C NMR spectrum of 1 revealed the presence of 21 carbons. Of these, 6 (δ_C 99.5, 75.2, 78.3, 71.8, 77.6, and 62.9) were assigned for a glucose unit, and the 15 remaining carbons were suggested for a sesquiterpene. Three double bonds [δ_C 115.8 (CH₂)/144.5 (CH), 126.4 (CH)/135.0 (C), and 126.1 (CH)/140.5 (CH)], 2 oxygenated quaternary carbons [δ_C 71.1 (C-11) and 81.4 (C-3)], 4 methyl groups [δ_C 16.1, 23.3, and 30.8 x2], and an anomeric carbon [δ_C 99.5 (C-1ʹ)] were identified (Table 1). The ¹H NMR spectrum of 1 also indicated 4 methyl groups [δ_H 1.40 and 1.60 (each 3H, s) and 1.28 (6H, s)], 6 olefinic protons of 3 double bonds and an anomeric proton (δ_H 4.38, d, J  =  7.5 Hz). The vinyl group was evident by signals at δ_H 5.21/5.25 (CH₂) and 5.95 (dd, J  =  17.5, 11.0 Hz, H-2), and a trans double bond was at δ_H 5.60 (dt, J  =  16.0, 6.0 Hz, H-9) and 5.60 (d, J  =  16.0 Hz). The NMR assignment of 1 was determined by analyzing the HSQC, COSY, and HMBC spectra. In the COSY spectrum, correlations of H-1/H-2, H-4/H-5/H-6, H-8/H-9/H-10, and H-1ʹ/H-2ʹ/H-3ʹ/H-4ʹ/H-5ʹ/H-6ʹ were clearly observed. In addition, correlations from H-1/H-2 to C-3, H-15 to C-2/C-3/C-4, H-14 to C-6/C-7/C-8, and H-12/H-13 to C-10/C-11 were observed in the HMBC spectrum, indicating that the aglycone of 1 was 3,11-dihydroxy-3,7,11-trimethyldodeca-16,9-triene (Table 1). ⁸ The HMBC interactions of H-1ʹ (δ_H 4.38)/C-3 (δ_C 81.4) confirmed the glucose moiety linked to C-3. A β-form of the glycosidic linkage was suggested from a large J value (7.5 Hz) of the H-1ʹ proton at δ_H 4.38. The E-geometry of the C-6/C-7 double bond was determined by the NOE cross-peaks between H₃-14 and H-5, and between H-6 and H-8 (Figure 2). Compound 1 was acid hydrolyzed to obtained D-glucose. ⁹ The positive specific rotation value of 1 suggested a (3S)-configuration.^10,11 To the best of our knowledge (from Sci. finder), all 3-hydroxy-3,7,11-trimethyldodeca-16,9-triene compounds have a (3S)-configuration. Thus, 1 was elucidated as (3S,6E,9E)-3-O-β-D-glucopyranosyl-11-hydroxy-3,7,11-trimethyldodeca-16,9-triene and named fissbalanoside A (Supplemental Figures S1 to S7).

Figure 1.

Structures of 1 and 2.

Figure 2.

Key HMBC and COSY correlations of 1 and 2.

Table 1.

NMR Spectroscopic Data for 1 and 2 in CD₃OD.

Pos.	aδ C	bδH (mult, J in Hz)	aδ C	bδH (mult, J in Hz)
1	115.8	5.21 (dd, 17.5, 1.0) 5.25 (dd, 11.0, 1.0)	115.7	5.20 (dd, 17.5, 1.0) 5.25 (dd, 11.0, 1.0)
2	144.5	5.95 (dd, 17.5, 11.0)	144.5	5.96 (dd, 17.5, 11.0)
3	81.4	–	81.4	–
4	42.6	1.64 (m)	42.5	1.62 (m)
5	23.6	2.07 (m)	23.6	2.06 (m)
6	126.4	5.19 (t, 7.0)	128.7	5.17 (t, 7.5)
7	135.0	–	132.7	–
8	43.5	2.68 (d, 6.0)	49.1	2.06 (m)/2.23 (m)
9	126.1	5.60 (dd, 16.0, 6.0)	68.0	4.43 (dt, 8.5, 7.0)
10	140.5	5.60 (16.0)	129.4	5.12 (d, 8.5)
11	71.1	–	134.8	–
12	30.8	1.28 (s)	25.9	1.72 (s)
13	23.3	1.40 (s)	18.3	1.67 (s)
14	16.1	1.60 (s)	16.6	1.64 (s)
15	30.8	1.28 (s)	23.3	1.40 (s)
1ʹ	99.5	4.38 (d, 7.5)	99.6	4.37 (d, 7.5)
2ʹ	75.2	3.16 (dd, 7.5, 8.8)	75.2	3.18 (dd, 7.5, 8.8)
3ʹ	78.3	3.34 (t, 8.8)	78.3	3.30 (t, 8.8)
4ʹ	71.8	3.29 (t, 8.8)	71.8	3.29 (t, 8.8)
5ʹ	77.6	3.19 (m)	77.6	3.19 (m)
6ʹ	62.9	3.83 (dd, 2.0, 11.6) 3.65 (dd, 5.5, 11.6)	62.8	3.83 (dd, 2.0, 11.6) 3.65 (dd, 5.5, 11.6)

^a 125 MHz.

^b

500 MHz.

The ¹³C and ¹H NMR data of 2 resembled those of 1 (Table 1) except that the signal for 1 oxygenated quaternary carbon at δ_C 71.1 (in 1) was replaced by the signal of 1 methine carbinol group at δ_C 68.0 (in 2). The HMBC correlations from H₃-12/H₃-13 to C-10/C-11 and from H-9 (δ_H 4.43) to C-11 were observed suggesting 1 double bond was located at C-10/C-11, and 1 hydroxy group at C-9. This was further confirmed by COSY cross peaks of H-8/H-9/H-10 (Figure 2). The glucose moiety linked to C-3 was similar to that of 1, as revealed from HMBC correlations of H-1ʹ (δ_H 4.37) and C-3 (δ_C 81.4). Therefore, the suggested molecular formula of 2 was C₂₁H₃₆O₇, which was further determined from HR-ESI-MS (found m/z 435.2150 [M  +  ³⁵Cl]⁻, calcd for [C₂₁H₃₆O₇³⁵Cl]⁻: 435.2155 and m/z 437.2131 [M  +  ³⁷Cl]⁻, calcd for [C₂₁H₃₆O₇³⁷Cl]⁻: 437.2125). The coupling constant observed for the anomeric proton at δ_H 4.37 (J  =  7.5 Hz) indicated that the glycoside linkage must be in the β-form. Compound 2 was acid hydrolyzed to obtained D-glucose. ⁹ Due to natural biosynthesis, the configuration at C-3 was suggested to be 3S.^10-12 However, the C-9 absolute configuration was not determined. From the above evidence, 2 was elucidated as (3S,6E)-3-O-β-D-glucopyranosyl-9-hydroxy-3,7,11-trimethyldodeca-1,6,10-triene and named fissbalanoside B (Supplemental Figures S8 to S13).

Sesquiterpenoids have been reported to have inhibitory effects on NO production. Therefore, compounds 1 and 2 were screened for their NO production inhibition in LPS-stimulated RAW 264.7 cells, as previously described.^13,14 At a concentration of 100 µM, these compounds did not significantly affect the growth of cells. Therefore, 1 and 2 were evaluated for their NO inhibitory effects at diluted concentrations ranging from 0.8 to 100 µM. Compounds 1 and 2 exhibited weak inhibition of NO production with IC₅₀ values of 52.12  ±  1.78 and 65.27  ±  1.82 µM, respectively, compared to that of the positive control, dexamethasone, IC₅₀: 14.20  ±  0.54 µM (Supplemental Table S1).

Materials and Methods

General

Refer to Supplemental Material

Plant Materials

F balansae branches were obtained from Me Linh, Vinh Phu, Vietnam and identified by Dr NT Cuong, IEBR, VAST. The voucher specimen (NCCT-P176) has been kept at IMBC.

Extraction and Isolation

Refer to Supplemental Material.

Fissbalanoside A (1)

Colorless powder, $[\alpha {]}_D^{25}$ :  +  13.0 (c 0.1, MeOH); HRESIMS m/z 435.2146 [M  +  ³⁵Cl]⁻, calcd for [C₂₁H₃₆O₇³⁵Cl]⁻: 435.2155 (Δ  =  -2.1 ppm); 437.2131 [M  +  ³⁷Cl]⁻, calcd for [C₂₁H₃₆O₇³⁷Cl]⁻: 437.2125 (Δ  =    +  1.4 ppm); ¹³C NMR and ¹H NMR data: see Table 1.

Fissbalanoside B (2)

Colorless powder, $[\alpha {]}_D^{25}$ :  +  7.9 (c 0.1, MeOH); HRESIMS m/z 435.2150 [M  +  ³⁵Cl]⁻, calcd for [C₂₁H₃₆O₇³⁵Cl]⁻: 435.2155 (Δ  =  -1.1 ppm); 437.2131 [M  +  ³⁷Cl]⁻, calcd for [C₂₁H₃₆O₇³⁷Cl]⁻: 437.2125 (Δ  =    +  1.4 ppm); ¹³C NMR and ¹H NMR and data: see Table 1.

Acid Hydrolysis

See supporting information. ⁹

Conclusions

From F balansae branches, 2 new sesquiterpene glycosides, fissbalanoside A and fissbalanoside B, were isolated. Their chemical structures were determined by analysis of their NMR and HR-ESI mass spectra.

Supplemental Material

sj-docx-1-npx-10.1177_1934578X231176161 - Supplemental material for Two New Sesquiterpene Glycosides from Fissistigma balansae Merr

Supplemental material, sj-docx-1-npx-10.1177_1934578X231176161 for Two New Sesquiterpene Glycosides from Fissistigma balansae Merr by Pham T Tham, Do T Trang, Bui H Tai and Phan V Kiem in Natural Product Communications