Three New Oleanane-Type Triterpenoidal Glycosides from Impatiens Balsamina and Their Biological Activity
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plants Article Three New Oleanane-Type Triterpenoidal Glycosides from Impatiens balsamina and Their Biological Activity Tae Hyun Lee 1, Won Se Suh 1, Lalita Subedi 2,3, Sun Yeou Kim 2,3 , Sang Un Choi 4, Kang Ro Lee 1 and Chung Sub Kim 1,* 1 School of Pharmacy, Sungkyunkwan University, Suwon 16419, Korea; [email protected] (T.H.L.); [email protected] (W.S.S.); [email protected] (K.R.L.) 2 Gachon Institute of Pharmaceutical Science, Gachon University, Incheon 21936, Korea; [email protected] (L.S.); [email protected] (S.Y.K.) 3 College of Pharmacy, Gachon University, Hambakmoero, Yeonsu-gu, Incheon 21936, Korea 4 Korea Research Institute of Chemical Technology, Daejeon 34114, Korea; [email protected] * Correspondence: [email protected]; Tel.: +82-31-290-7727 Received: 22 July 2020; Accepted: 19 August 2020; Published: 24 August 2020 Abstract: Three new oleanane-type triterpenoidal glycosides, imbalosides A–C (1–3), were isolated from the white flowers of Impatiens balsamina. The structures of these phytochemical constituents (1–3) were elucidated through 1D and 2D Nuclear Magnetic Resonance (NMR) and Mass Spectrometry (MS) data analyses followed by chemical methods. All the characterized compounds (1–3) were evaluated for their antiproliferative activity against four human tumor cell lines (A549, SK-OV-3, SK-MEL-2, and BT549) and their anti-neuroinflammatory activity on the basis of inhibition levels of nitric oxide (NO) in the lipopolysaccharide (LPS)-stimulated murine microglia BV-2 cell lines. Keywords: Impatiens balsamina; balsaminaceae; triterpenoidal glycosides; cytotoxicity; anti-neuroinflammation 1. Introduction Impatiens balsamina L., known as garden balsam or rose balsam, is an annual plant belonging to the family Balsaminaceae and is widely distributed in Korea, Japan, and mainland China. Diverse parts of I. balsamina, including flowers, stems, and leaves, have long been used as traditional medicines to treat various diseases. The flowers of I. balsamina have been used as remedies for lumbago, burns, and scalds [1], whereas its aerial parts have been used to treat articular rheumatism, abscesses, and tumors [2]. In the previous research on this plant, 1,4-naphthoquinone derivatives showed a variety of pharmacological effects such as antitumor, anti-inflammatory, and hepatoprotective activities [3–5]. As part of the continuing studies to identify the bioactive constituents from Korean medicinal plants [6–11], we previously conducted a phytochemical investigation on the MeOH extract of the white flowers of I. balsamina, which led to the isolation and characterization of phenolic compounds including mono- and biflavonoids with cytotoxic, anti-inflammatory, and neuroprotective activities [12,13]. In order to discover bioactive molecules in other structural classes from this plant, we further investigated its EtOAc-soluble layer and identified three new oleanane-type triterpenoidal glycosides (1–3) (Figure1). The chemical structures of the new compounds ( 1–3) were established on the basis of spectroscopic (1D and 2D NMR) and spectrometric [High Resolution Fast Atom Bombardment MS (HRFABMS)] analyses as well as chemical methods. The isolates (1–3) were tested for their cytotoxicity against four human tumor cell lines and anti-neuroinflammatory activity using lipopolysaccharide (LPS)-stimulated murine microglia BV-2 cell lines. Plants 2020, 9, 1083; doi:10.3390/plants9091083 www.mdpi.com/journal/plants Plants 2020, 9, x FOR PEER REVIEW 2 of 9 Plants 2020, 9, x FOR PEER REVIEW 2 of 9 cytotoxicity against four human tumor cell lines and anti-neuroinflammatory activity using cytotoxicitylipopolysaccharide against (LPS)-stimulated four human tumor murine cell microglia lines andBV-2 anti-neuroinflammatorycell lines. activity using Plants 2020, 9, 1083 2 of 9 lipopolysaccharide (LPS)-stimulated murine microglia BV-2 cell lines. Figure 1. Chemical structures of compounds 1–3. Figure 1. Chemical structures of compounds 1–3. Figure 1. Chemical structures of compounds 1–3. 2. Results and Discussion 2. Results and Discussion 2. Results and Discussion 2.1. Structure Elucidation 2.1. Structure Elucidation Imbaloside A A ( (11)) was was isolated isolated as as a a colorless colorless gum. gum. The The molecular molecular formula formula of of1 was1 was determined determined as C41H66O14 based on the [M – H]– ion peak– at m/z 781.4368 (calcd for C41H65O14–, 781.4380,– error = 1.5 as C41ImbalosideH66O14 based A (1) onwas the isolated [M – as H] a colorlession peak gum. at m The/z 781.4368 molecular (calcd formula for of C 411 Hwas65O determined14 , 781.4380, as 1 ppm) from the HRFABMS analysis.– The H NMR spectrum1 of 1 showed a broad– singlet at δH 5.30 for errorC41H66=O1.514 based ppm )on from the the [M HRFABMS – H] ion peak analysis. at m/z The 781.4368H NMR (calcd spectrum for C41 ofH165showedO14 , 781.4380, a broad error singlet = 1.5 at an olefinic proton, overlapped signals 1 from δH 3.20 to 4.38 for oxygenated methine/methylene δppm)H 5.30 from for an the olefinic HRFABMS proton, analysis. overlapped The signalsH NMR from spectrumδH 3.20 of to 4.381 showed for oxygenated a broad singlet methine at/ methyleneδH 5.30 for protons, seven singlets at δH 1.40, 1.09, 1.05, 1.00, 0.97, 0.93, and 0.88 for methyl protons, and the others protons,an olefinic seven proton, singlets overlapped at δH 1.40, signals 1.09, 1.05,from 1.00,δH 3.20 0.97, to 0.93, 4.38 and for 0.88oxygenated for methyl methine/methylene protons, and the in the region from δH 2.28 to 0.82. Among a total of 41 carbons present in this molecule, 40 resonances othersprotons, in seven the region singlets from at δHδ 1.40,H 2.28 1.09, to 1.05, 0.82. 1.00, Among 0.97, 0.93, a total and of 0.88 41 carbonsfor methyl present protons, in thisand the molecule, others were observed in the 13C NMR13 spectrum of 1, including 30 peaks for typical oleanane-type 40in the resonances region from were δ observedH 2.28 to in0.82. the AmongC NMR a total spectrum of 41 ofcarbons1, including present 30 in peaks this formolecule, typical oleanane-type40 resonances triterpenoids [14–18] with13 two olefinic carbons at δC 144.6 and 124.2 and four oxygenated carbons at triterpenoidswere observed [14 –in18 the] with C two NMR olefinic spectrum carbons of at 1δ,C including144.6 and 124.230 peaks and fourfor oxygenatedtypical oleanane-type carbons at δC 91.1, 78.8, 70.4, and 65.8, and ten peaks for β-glucuronic acid (δC 106.9, 75.6, 78.1, 73.8, and 76.8) δtriterpenoidsC 91.1, 78.8, 70.4,[14–18] and with 65.8, two and olefinic ten peaks carbons for β at-glucuronic δC 144.6 and acid 124.2 (δC and106.9, four 75.6, oxygenated 78.1, 73.8, carbons and 76.8) at and β-xylopyranose (δC 102.8, 74.5, 77.0, 71.2 and 66.5). These 11H and 13C NMR data implied that 1 is andδC 91.1,β-xylopyranose 78.8, 70.4, and (δC 65.8,102.8, and 74.5, ten 77.0, peaks 71.2 for and β-glucuronic 66.5). These acidH and(δC 106.9,C NMR 75.6, data 78.1, implied 73.8, and that 76.8)1 is anand oleanane-type β-xylopyranose triterpenoidal (δC 102.8, 74.5, glycoside 77.0, 71.2 with and two 66.5). sugar These moieties, 1H and and 13C NMR its core data structure implied including that 1 is thean oleanane-type location location of of a a double doubletriterpenoidal bond bond and andglycoside four four oxygenate oxygenated with twod carbonssugar carbons moieties, was was established establishedand its core through throughstructure analysis analysis including of the of theDistortionless Distortionlesslocation of aEnhancement double Enhancement bond byand byPolarization four Polarization oxygenate Transfer Transferd carbons (DEPT), (DEPT), was establishedcorrelation correlation throughspectroscopy spectroscopy analysis (COSY), of the HeteronuclearDistortionless Enhancement Single Quantum by CorrelationPolarization (HSQC),(HSQC) Transfer, and (DEPT), Heteronuclear correlation Multiple spectroscopy Bond Correlation (COSY), (HMBC)Heteronuclear spectraspectra Single (Figure(Figure Quantum2 2,, Supplementary Supplementary Correlation Materials). materials). (HSQC), and Heteronuclear Multiple Bond Correlation (HMBC) spectra (Figure 2, Supplementary materials). Figure 2. Key COSY (blue bold) and HMBC (red arrows) correlations of 1–3. Figure 2. Key COSY (blue bold) and HMBC (red arrows) correlations of 1–3. The β-configurationsFigure 2. Key of COSY the two(blue anomeric bold) and HMBC carbons (red on arrows) the glucuronic correlations acid of 1 and–3. xylopyranose wereThe assigned β-configurations by the relatively of the two large anomeric3J coupling carbons constants on the (7.1glucuronic Hz) between acid and H-1 xylopyranose and H-2 of were both 3 sugars.assignedThe These βby-configurations the two relatively sugar of unitslarge the weretwoJ coupling anomeric confirmed constants carbons to be (7.1 on connected the Hz) glucuronic between at C-3 H-1 acid (glucuronic and and H-2 xylopyranose acid)of both and sugars. were C-22 These two sugar units were 3confirmed to be connected at C-3 (glucuronic acid) and C-22 (xylopyranose)assigned by the byrelatively observing large HMBC J coupling cross-peaks constants of H-3 (7.1/C-1 Hz)0 and between H-22/ C-1H-100 and(Figure H-22 of). Theboth relative sugars. configurationThese(xylopyranose) two sugar at by C-3 observingunits was confirmedwere HMBC confirmed by cross-peaks the strongto be nuclearof connected H-3/C-1 Overhauser′ andat C-3H-22/C-1 eff(glucuronicect′′ (NOE) (Figure correlationacid) 2). The and relative of C-22 H-3 with(xylopyranose)configuration H-5 along at by withC-3 observing was the confirmed mild HMBC correlation by cross-peaks the ofstrong H-3 ofnuclear with H-3/C-1 H-1ax Overhauser′ and (Figure H-22/C-13 effectA).