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. 2023 Apr 6;8(15):14240–14246. doi: 10.1021/acsomega.3c01247

Synthesis, Structure Revision, and Anti-inflammatory Activity Investigation of Putative Blumeatin

Kai Xia †, Wei-Jin Qi †, Xiao-Qiang Wu ∥, Yu-Yang Song †, Jun-Jie Zhu †, Yi Ai †, Zhen Cui †, Zheng-Ping Zhang †, Shu-Ai Tang †, Yu-Ting Gui †, Yue Yuan †, Lu Wang §, Hang Zhong †,‡,*
PMCID: PMC10116622  PMID: 37091405

Abstract

graphic file with name ao3c01247_0008.jpg

Blumeatin, reported herein, bearing two hydroxyl groups at C3′ and C5′ of ring B, is isolated from the traditional Chinese medicine Blumea balsamifera. But the isolation procedure of blumeatin from plants has limitations of prolonged duration and high cost. A procedure featuring Lewis acid-catalyzed ring closure and chiral resolution via Schiff base intermediates is provided here to prepare optically pure blumeatin and its R-isomer efficiently. Furthermore, the structure revision of putative blumeatin based on a logically synthetic procedure and NMR spectroscopic analysis was conducted. The 1D and 2D NMR data analysis unambiguously confirmed our proposal that the reported blumeatin structure has been misassigned as it corresponds to sterubin, which contains two hydroxyl groups at C3′ and C4′ of ring B. Finally, the results of the ear-swelling test exhibited that synthetic (±)-blumeatin and (±)-sterubin had moderate anti-inflammatory activity which was less than that of (−)-sterubin.

1. Introduction

Blumea balsamifera is a renowned medicinal plant in the Chinese Pharmacopoeia.1−5 It is rich in chemical components such as flavonoids, sesquiterpenes, volatile oils, and sterols.6,7 Blumeatin (Figure 1) is one of the flavanones with bioactivities extracted from B. balsamifera. The structure of blumeatin was reported to be (S)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one in the literature.1,5,8−13 Particularly, blumeatin exhibited significant anti-inflammatory activity and promoted fat cell differentiation.14,15 Our previous research indicated that the anti-inflammatory activity of blumeatin was slightly less than that of dexamethasone in an in vitro experiment.16 However, isolation from B. balsamifera represents the main way to obtain blumeatin currently, which has low efficiency and high cost. Therefore, establishing a synthetic approach to obtain blumeatin could have an important significance in the use of blumeatin.

Figure 1.

Figure 1

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Structure of putative blumeatin.

There are many ways to chemically synthesize flavonoids. Patel and Shah17 reported a one-step procedure for flavone preparation via an iodine-mediated oxidative ring-closure reaction with chalcone. However, acquisition of chalcone precursors waas not very convenient, and the yields of the ring-closure reactions were not high enough. A β-diketone formed by the Baker–Venkataraman method18,19 suffered from an acid-catalyzed ring-closure reaction to afford flavones. But this procedure was not environmentally friendly due to the large amount of waste acid produced. Applications of transition-metal catalysts could efficiently construct flavones,20,21 but they were costly and tough operating conditions were required. Zhang’s group22 provided an approach for the preparation of flavones: a ring-closure reaction from substituted acetophenones and substituted benzaldehydes in the presence of boric acid and piperidine to generate flavones. This procedure was simple, efficient, and easy to operate. Therefore, we planned to prepare blumeatin based on Zhang’s method.

We herein report a synthetic procedure to obtain blumeatin (Figure 2) involving the methylation of (2,4,6-trihydroxy-phenyl)ethan-1-one, construction of the flavanone nucleus under Lewis acid catalysis, and chiral resolution via a Schiff base to produce optically active blumeatin.

Figure 2.

Figure 2

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Synthetic route of (−)-blumeatin.

2. Results and Discussion

2.1. Synthesis of Optically Active (−)-Blumeatin and (+)-Blumeatin

The key step to construct blumeatin was the closure of the C-ring. It could be completed by the reaction of 3,5-dihydroxybenzaldehyde and 1-(2,6-dihydroxy-4-methoxyphenyl) ethan-1-one in the presence of boric acid, piperidine, and silica gel. Then, (±)-blumeatin (1) was obtained with a 75% total yield (Scheme 1). Fortunately, followed by imidization of 4-carbonyl of (±)-blumeatin with an optically pure (R)-1-phenylethan-1-amine, the diastereomeric imine intermediates could be separated favorably by silica column chromatography to give the same amount of optically active 2A and 2B. Finally, an acid-mediated hydrolysis produced optically pure (+)-blumeatin (1A) or (−)-blumeatin (1B), respectively.

Scheme 1. Syntheses of (+)-Blumeatin and (−)-Blumeatin.

Scheme 1

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2.2. Electronic Circular Dichroism Analysis of 1A and 1B

To clarify the absolute configurations of 1A and 1B, the electronic circular dichroism (ECD) spectra of 1A and 1B were tested in MeOH (0.2668 mg/mL and 0.2735 mg/mL, respectively) in the 200–400 nm range. In the ECD spectrum of 1A, a negative cotton effect is visible at 280 nm (Figure 3). For 1B, a positive cotton effect is observed at 290 nm. Simultaneously, ECD calculations were performed by the time-dependent density functional theory-predicted curve calculated at the quantum mechanical level. The calculated ECD spectra of 1A and 1B were consistent with their experimental counterparts. According to the ECD spectra rule of flavanones reported by Gaffield,231A was assigned as an R-configuration and 1B was an S-isomer.

Figure 3.

Figure 3

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ECD spectra of 1A and 1B.

2.3. Structure Revision of Putative Blumeatin

According to HRMS, ECD, 1H NMR, 13C NMR, and 2D-NMR data, combined with the watertight procedure and synthetic raw materials, the target molecules 1A and 1B were determined as (R)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxy- chroman-4-one and (S)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one, respectively. Intriguingly, both the synthetic compounds 1A and 1B could not match with the natural blumeatin (4A) isolated by our group and others (purchased from Tauto Biotech, Co., Ltd. or Rhawn, Co., Ltd.) in TLC (Figure S37) or HPLC experiments (Figures S1, S8, S13, and S34). Further structural analysis of the natural “blumeatin” (4A) was pressing.

2.3.1. 1: Synthetic Blumeatin; 4A: Putative Blumeatin

According to 1H NMR, 13C NMR and HMBC data of racemic compound 1 (Table 1, Figure S5), signals at δ H/C 5.45 (H-2)/78.9 (C-2), 3.16 & 2.77 (H-3)/42.7 (C-3), 197.0 (C-4), 102.9 (C-4a), 12.09 (OH-5)/163.6 (C-5), 6.09 (H-6)/91.6 (C-6), 3.80 (OCH3-7)/167.9 (C-7), 6.13 (H-2)/94.3 (C-8), 163.1 (C-8a), 141.0 (C-1′), 104.9 (C-2′, C-6′), 158.9 (C-3′, C-5′), 103.1 (C-4′), were consistent with the scaffold of 5-hydroxy-7-methoxychroman-4-one. In addition, two symmetrical signals of 1H NMR at 6.34 ppm (H-2′, H-6′), 9.37 ppm (OH-3′, OH-5′) together with a triplet at 6.21 (J = 2.1 Hz, remote coupling with H-2′ and H-6′, respectively), were corresponding to two hydroxy groups at C3′ and C5′ of ring B, respectively.

Table 1. 1H NMR, 13C NMR, and HMBC Data for 1 and 4A in DMSO-d6.
  compound 1
 compound 4A
position δHa δCa HMBCa,b δHa δCa HMBCa,b
C2 5.45, dd (12.1, 3.2) 78.9, CH 1′, 2′, 6′, 4, 8a 5.45, dd (12.6, 3.0) 79.1, CH 1′, 2′, 5′, 6′
C3 3.16, dd (17.2, 12.1); 2.77, dd (17.2, 3.2) 42.7, CH2 1′, 2, 4, 4a 3.22, dd (17.1, 12.6); 2.72, dd (17.1, 3.0) 42.6, CH2 1, 2, 4, 4a
C4   197.1, C     197.4, C  
C4a   102.9, C     103.1, C  
C5   163.7, C     163.6, C  
C6 6.09, d (2.3) 95.1, CH 5, 7, 8, 8a, 4a 6.08, d (2.2) 96.0, CH 5, 7, 8, 8a, 4a
C7   167.9, C     167.9, C  
C8 6.13, d (2.3) 94.4, CH 5, 7, 8, 8a, 4a 6.10, d (2.2) 94.2, CH 5, 7, 8, 8a, 4a
C8a   163.1, C     163.3, C  
C1′   141.0, C     129.7, C  
C2′ 6.34, d (2.0) 104.9, CH 2, 3′, 4′, 5′, 6′ 6.89, s 114.8, CH 1′, 2, 3′, 4′, 5′
C3′   158.9, C     145.6, C  
C4′ 6.21, t (2.1, 2.1) 103.1, CH 2′, 3′, 5′, 6′   146.2, CH  
C5′   158.9, C   6.73–6.78c, m 115.8, CH 1′, 2′, 2, 3′, 4′
C6′ 6.34, d (2.0) 104.9, CH 2, 2′, 3′, 4′, 5′ 6.73–6.78c, m 118.5, CH 1′, 2, 3′, 4′
C3′-OH 9.37, s     9.09, s    
C4′-OH       9.05, s    
C5′-OH 9.37, s          
C7–OCH3 3.80, s 56.4, CH3   3.79, s 56.6, CH3  
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a

Measured at 400 MHz (25 °C).

b

From proton to stated carbon(s).

c

Signal overlapped.

Similarly, compound 4A also had a skeleton of 5-hydroxy-7-methoxychroman-4-one (Table 1). However, 13C NMR signals at 129.7 (C-1′), 114.8 (C-2′), 145.6 (C-3′), 146.2 (C-4′), 115.8 (C-5′), and 118.5 (C-6′) indicated that there were no two carbon atoms with identical chemical environment in the B-ring. Together with 1H NMR signals at 6.89 (H-2′), 9.09 (OH-3′), 9.05 (OH-4′), 6.73–6.78 (H-5′ and H-6′ overlapped accidently), it indicated that two hydroxyl groups were severally located at the C3′ and C4′ in the B-ring of 4A.

The positions of hydroxy groups in the B-ring of 1 or 4A could be further validated by HMBC data (Table 1). For compound 4A, cross-peaks between C2 and H2′, H6′, H5′ were readily observed, while C2 was just correlated with H2′ and H6′ in the HMBC spectrum of compound 1, for both C3′ and C5′ were substituted with hydroxy groups, and we also could not see the correlation from H4′ to C2 in HMBC. Taken together with the results of HMBC, 1 could be validated as 2-(3,5-dihydroxyphenyl)-5- hydroxy-7-methoxychroman-4-one and 4A was determined as 2-(3,4-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one (Figure 4).

Figure 4.

Figure 4

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Structural comparison of 1 and 4A.

The structure of 4A coincided with the structure of sterubin, namely, (S)-2-(3,4-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one.24 If the reactant of 3,5-dihydroxybenzaldehyde in our synthetic procedure was replaced with 3,4-dihydroxybenzaldehyde, the (±)-2-(3,4-dihydroxyphenyl)-5-hydroxy-7-methoxy-chroman-4-one (4, (±)-sterubin) could be obtained (Figure 5). Its 1H NMR and 13C NMR data were highly consistent with those of 4, so was those of mentioned in literature.24−28

Figure 5.

Figure 5

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Synthesis of compound racemic 4.

In addition, the melting points of newly synthetic compounds were measured many times on two micro melting point apparatuses, and the average values were taken to determine that the melting point of racemate 1 was 181–183 °C. Similarly, the melting points of optically active 1A and 1B were 180–181 °C and 179–181 °C, respectively. While the earliest reported melting point of the putative blumeatin was 221–223 °C8 or 218–220 °C in other literature,5,29 which were neither consistent with that of 1 nor those of 1A and 1B. On the other hand, the melting point of compound 4 was 217–219 °C after testing, which did match with the data reported in the literature.24 Taken together the consistent results of NMR and HPLC between 4 and 4A (Figures S8–S19), we can unambiguously propose that the structure of the putative blumeatin is corresponding to that of sterubin (mp 215–216 °C).24

2.4. Anti-inflammatory Activity of Synthetic Blumeatin

To investigate the anti-inflammatory activities of our synthesized compounds, we performed ear-swelling experiments on mice, with dexamethasone (10 mg/kg) as a positive control, and the results are summarized in Table 2. It could be seen that the preventative anti-inflammatory activity of isolated blumeatin (sterubin) 4A presented dose-dependent. The swelling inhibitory rate of 4A at high dose (30 mg/kg) was 73.53%, which was about 1/3 as potent as to positive control (78.39% at 10 mg/kg), and the anti-inflammatory activities of 4A were also reported in other works.28,30,31 Similarly, both 1 and 4 inhibited ear swelling in a dose-dependent manner, but 4 was slightly more potent than 1. It indicated that 3,4-dihydroxy in the B-ring was superior to its 3,5-dihydroxy counterpart. Simultaneously, racemic 4 was slightly less potent than S-isomer 4A, suggesting S-isomer might be the eutomer on anti-inflammatory activity.

Table 2. Comparison of Swelling Degree of Mice in Each Groupa.

testing sets dose (mg/kg) swelling (mg) inhibition rate (%)
blank set   8.41 ± 2.28  
solvent set   7.23 ± 1.99  
dexamethasone 10 1.57 ± 1.00** 78.30
1 7.5 4.47 ± 2.78** 38.17
  15 4.29 ± 2.41** 40.66
  30 3.71 ± 1.22** 48.70
4A 7.5 5.23 ± 2.11* 27.74
  15 2.45 ± 2.17** 66.13
  30 1.92 ± 0.85** 73.53
4 7.5 6.58 ± 2.45** 9.79
  15 5.36 ± 2.20** 26.58
  30 3.32 ± 1.60** 54.52
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a

* P < 0.05, **P < 0.01. 1: (±)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one; 4A: putative blumeatin; 4: (±)-2-(3,4-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one.

3. Conclusions

The structure of putative blumeatin was proposed as (S)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one by its discoverer. We here provide a synthetic and resolving procedure for efficiently forming optically pure blumeatin. However, the synthesized blumeatin was not consistent with its isolated counterpart (commercially available “blumeatin”). The data of 1H NMR and 13C NMR, HMBC, and HPLC and the results of our synthetic procedure supported that the isolated blumeatin match with sterubin [(S)-2-(3,4-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one]. The results of the ear-swelling test exhibited that synthetic (±)-blumeatin and (±)-sterubin had moderate anti-inflammatory activity, which was less than that of putative blumeatin (sterubin).

4. Experimental Section

4.1. Reagents and Instruments

Partial reactions were carried out in flame-dried glassware under a nitrogen atmosphere. Reagents used were purchased from Aladdin, Macklin, Innochem, or TLC unless otherwise noted. Chromatographic separation used silica gel, AR, 200–300 mesh. 1H and 13C NMR spectra were recorded on a Bruker (Avance) 400 MHz NMR instrument using CDCl3 and DMSO-d6 as solvents. Infrared spectra were acquired with KBr particles on a Shimadzu FT-IR-8400S spectrometer. Optical rotations were determined on an Insmark digital polarimeter using a sodium (589 nm, D-line) lamp and reported as follows: c = 0.20 g/100 mL, ethanol, 20 °C. The melting point of the compound was recorded by the X-4D micro melting point apparatus. An Applied Photophysics Chirascan spectrometer was employed to get ECD spectra. Retention time was obtained in Agilent 1260 HPLC. TLC analysis was visualized using UV, vanillin, and iodine elemental staining. High-resolution mass spectra were obtained using AB SCIEX X500R QTOF.

4.2. Synthetic Procedures

4.2.1. 2,6-Dihydroxy-4-methoxy-acetophenone (3)

2,4,6-Trihydroxy acetophenone (8 g, 0.048 mol, 1 equiv) and anhydrous potassium carbonate (13.14 g, 0.095 mol, 2 equiv) were dissolved in acetone (100 mL). Dimethyl sulfate (2.7 mL, 0.029 mol, 0.6 equiv) was added dropwise, and the reaction mixture was stirred at 60 °C under nitrogen for 24 h. Then, the mixture was allowed to slowly cool to room temperature. After the reaction was quenched with 50 mL of water and diluted with 50 mL of ethyl acetate (EA). The layers were separated, and the aqueous layer was extracted 2× with 50 mL of EA. The combined organics were washed with brine and dried over Na2SO4. Finally, the evaporated residue was purified by column chromatography (3–4% EA in petroleum ether) to 2,6-dihydroxy -4-methoxy- acetophenone (3) (3.33 g, 35%) as a yellow solid. 1H NMR (400 MHz, CDCl3): δ 5.93 (s, 2H), 3.79 (s, 3H), 2.67 (s, 3H); mp 126–130 °C; HRMS (ESI/TOF) m/z calcd for C8H9O4, 183.06573; found, 183.06592 [M + H]+.

4.2.2. 2-(3,5-Dihydroxy phenyl)-5-hydroxy-7-methoxychromanone (1)

2,6-Dihydroxy-4-methoxy-acetophenone (3) (500 mg, 2.75 mmol, 0.8 equiv) and 3,5-dihydroxybenzaldehyde (470 mg, 3.41 mmol, 1 equiv) were added to a mixture of H3BO3 (337 mg, 5.41 mmol, 1.5 equiv) and silica gel (1.5 g) in DMF (30 mL), and then piperidine (113 μL, 1.4 mmol, 0.25 equiv) was added dropwise under a nitrogen atmosphere. The reaction mixture was stirred and heated at 120 °C under nitrogen for 10–12 h, followed by cooling to room temperature and diluting with acetone (50 mL) and filtering. The silica gel was rinsed with acetone (5 mL) three times. The acetone solution was combined with the filtrate and evaporated at reduced pressure to dryness. The residue was purified by column chromatography (12–16% EA in petroleum ether) to 2-(3,5-dihydroxy phenyl)-5-hydroxy-7-methoxychromanone (1) (750 mg, 75%) as a pale yellow solid. IR: 3290, 3140, 2992, 1603, 1354, 1263, 1148, 1090, 974, 825, 709; 1H NMR (400 MHz, DMSO-d6): δ 12.09 (s, 1H),9.37 (s, 2H), 6.34 (d, J = 2.0 Hz, 2H), 6.21 (t, J = 2.1, 2.1 Hz, 1H), 6.13 (d, J = 2.3 Hz, 1H), 6.09 (d, J = 2.3 Hz, 1H), 5.45 (dd, J = 12.1, 3.2 Hz, 1H),3.80 (s, 3H), 3.16 (dd, J = 17.2, 12.1 Hz, 1H), 2.77 (dd, J = 17.2, 3.2 Hz, 1H); 13C NMR (100 MHz, DMSO-d6): δ 197.1 (C4), 167.9 (C7), 163.7 (C5), 163.1 (C9), 158.9 (C3′ and C5′), 141.0 (C1′), 104.9 (C6′), 103.1 (C2′), 102.9 (C4′), 95.1 (C6), 94.4 (C8), 78.9 (C2), 56.4 (7-OCH3), 42.7 (C3); mp 181–183 °C; HRMS (ESI/TOF) m/z calcd for C16H14O6Na, 325.06881; found, 325.06749 [M + Na]+.

4.2.3. 5-((R)-5-Hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benzene-1,3-diol (2A)

2-(3,5-Dihydroxy phenyl)-5-hydroxy-7-methoxychromanone (1) (500 mg, 1.65 mmol, 1 equiv) and anhydrous potassium carbonate (450 mg, 3.26 mmol, 2 equiv) were dissolved in absolute ethanol (10 mL). (S)-(−)-α-methyl benzylamine (840 μL, 6.72 mmol, 4 equiv) was added dropwise, and the reaction mixture was stirred under reflux for 3.5 h. Then the mixture was allowed to slowly cooled to room temperature. After the reaction mixture was quenched with 30 mL of water, the aqueous layer was extracted 3× with 30 mL of EA. The combined organics were washed with brine, dried over Na2SO4 and evaporated to dryness and purified by column chromatography (40–50% EA in petroleum ether) to afford 5-((R)-5- hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benzene-1,3-diol (2A) (300 mg, 50%) as a pale yellow solid. 1H NMR (400 MHz, DMSO-d6): δ 9.34 (s, 2H), 7.48–7.28 (m, 5H), 6.33, (d, J = 1.9 Hz, 2H), 6.21 (t, J = 1.8 Hz, 1H), 5.84 (d, J = 2.4 Hz, 1H), 5.80 (d, J = 2.4 Hz, 1H), 5.04 (dd, J = 11.3, 3.2 Hz, 1H), 3.71 (s, 3H), 3.18 (dd, J = 16.6, 3.2 Hz, 1H), 3.07 (dd, J = 17, 11.3 Hz, 1H), 1.45 (m, 3H); mp 237–239 °C; HRMS (ESI/TOF) m/z calcd for C24H24NO5, 406.16545; found, 406.16468 [M + H]+.

4.2.4. 5-((S)-5-Hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benz-ene-1,3-diol (2B)

5-((S)-5-Hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benz-ene-1,3-diol (2B) was obtained by the same process of 2A (300 mg, 47%) as a pale yellow solid. 1H NMR (400 MHz, DMSO-d6): δ 9.34 (s, 2H), 7.48–7.28 (m, 5H), 6.33 (d, J = 2.1 Hz, 2H), 6.21 (t, J = 1.9 Hz, 1H), 5.84 (d, J = 2.4 Hz, 1H), 5.80 (d, J = 2.4 Hz, 1H), 5.08 (dd, J = 11.1, 3.1 Hz, 1H), 3.71 (s, 3H), 3.18 (dd, J = 17.4, 3.1 Hz, 1H), 3.07 (dd, J = 17.4, 11.1 Hz, 1H), 1.45 (m, 3H); mp 238–240 °C; HRMS (ESI/TOF) m/z calcd for C24H24NO5, 406.16545; found, 406.16471 [M + H]+.

4.2.5. (R)-2-(3,5-Dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one (1A)

5-((R)-5-Hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benz-ene-1,3-diol (2A) (200 mg, 0.48 mmol, 1 equiv) was dissolved in EA (10 mL), and 5 N hydrochloric acid was added (15 mL). The reaction mixture was stirred at 100 °C under nitrogen for 10 h. and the mixture was allowed to slowly cooled to room temperature. Then the reaction mixture was neutralized with sodium hydroxide solution and diluted with 20 mL of EA. The layers were separated, and the aqueous layer was extracted 2× with 10 mL of EA. The combined organics were washed with brine, dried over Na2SO4, and purified by column chromatography (12–16% EA in petroleum ether) to (R)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one (1A) (160 mg, 95%) as a yellow solid. IR: 3300, 3110,2890, 1600, 1420, 1350, 1260, 1200, 1144, 980, 820, 710; 1H NMR (400 MHz, DMSO-d6): δ 12.09 (s, 1H), 9.37 (s, 2H), 6.34 (d, J = 2.0 Hz, 2H), 6.21 (t, J = 2.1, 2.1 Hz, 1H), 6.13 (d, J = 2.3 Hz, 1H), 6.09 (d, J = 2.3 Hz, 1H), 5.45 (dd, J = 12.1, 3.2 Hz, 1H), 3.80 (s, 3H), 3.16 (dd, J = 17.2, 12.1 Hz, 1H), 2.77 (dd, J = 17.2, 3.2 Hz, 1H); 13C NMR (100 MHz, DMSO-d6): δ 197.1 (C4), 167.9 (C7), 163.7 (C5), 163.1 (C9), 158.9 (C3′ and C5′), 141.03 (C1′), 104.9 (C6′), 103.1 (C2′), 102.9 (C4′), 95.1 (C6), 94.4 (C8), 78.9 (C2), 56.4 (7-OCH3), 42.7 (C3); mp 180–181 °C; [α]Dt = +7.2 HRMS (ESI/TOF) m/z calcd for C16H14O6Na, 325.06881; found, 325.06790 [M + Na]+.

4.2.6. (S)-2-(3,5-Dihydroxyphenyl)-5-hydroxy-7-methoxychroman-4-one (1B)

5-((S)-5-Hydroxy-7-methoxy-4-(((R)-1-phenylethyl)imino)chroman-2-yl)benz-ene-1,3-diol (2B) was hydrolyzed by the same process of 2A to afford (S)-2-(3,5-dihydroxyphenyl)-5-hydroxy-7- methoxychroman-4-one 1B (150 mg, 90%) as a yellow solid. IR: 3340, 3163, 2868, 1596, 1430, 1301, 1255, 1190, 1144, 1071, 978, 822, 711; 1H NMR (400 MHz, DMSO-d6): δ 12.09 (s, 1H), 9.37 (s, 2H), 6.34 (d, J = 2.0 Hz, 2H), 6.21 (t, J = 2.1, 2.1 Hz, 1H), 6.13 (d, J = 2.3 Hz, 1H), 6.09 (d, J = 2.3 Hz, 1H),5.45 (dd, J = 12.1, 3.2 Hz, 1H), 3.80 (s, 3H), 3.16 (dd, J = 17.2, 12.1 Hz, 1H), 2.77 (dd, J = 17.2, 3.2 Hz, 1H); 13C NMR (100 MHz, DMSO-d6): δ 197.1 (C4), 167.9 (C7), 163.7 (C5), 163.1 (C9), 158.9 (C3′ and C5′), 141.03 (C1′), 104.9 (C6′), 103.1 (C2′), 102.9 (C4′), 95.1 (C6), 94.4 (C8), 78.9 (C2), 56.4 (7-OCH3), 42.7 (C3); mp 179–181 °C; [α]Dt = −7.0; HRMS (ESI/TOF) m/z calcd for C16H14O6Na, 325.06881; found, 325.06824 [M + Na]+.

4.2.7. 2-(3,4-Dihydroxy phenyl)-5-hydroxy-7-methoxychromanone (4)

Compound 4 (600 mg, 65%) as a white solid was obtained according to the synthetic method of compound 1. IR: 3331, 3116, 1594, 1437, 1354, 1247, 1189, 1148, 1065, 949, 817, 709, 544; 1H NMR (400 MHz, DMSO-d6): δ 12.1 (s, 1H), 9.16 (s, −OH), 9.08 (s, −OH), 6.85 (d, J = 1.28 Hz, 1H), 6.73–6.78 (m, 2H), 6.10 (d, J = 2.3 Hz, 1H), 6.08 (d, J = 2.3 Hz, 1H), 5.42 (dd, J = 12.6, 3.0 Hz, 1H), 3.79 (s, 3H), 3.24 (dd, J = 17.2, 12.6 Hz), 2.72 (dd, J = 17.2, 3.0 Hz); 13C NMR (100 MHz, DMSO-d6): δ 197.4 (C4), 167.9 (C7), 163.6 (C5), 163.3 (C9), 145.7 (C3′), 145.2 (C4′), 129.7 (C1′), 118.4 (C6′), 115.9 (C5′), 114.9 (C2′), 103.1 (C10), 95.0 (C6), 94.2 (C8), 79.1 (C2), 56.4 (7-OCH3), 42.6 (C3); mp 217–219 °C; HRMS (ESI/TOF) m/z calcd for C16H14O6Na, 325.06881; found, 325.06784 [M + Na]+.

4.3. Ear Swelling Test on Mice

The Animal Care Welfare Committee of Guizhou University gave the approval to conduct animal experiments. One hundred and twenty Kunming mice were randomly divided into 12 groups (with 10 mice in each group), comprising a blank group (normal saline), a solvent group (Vethanol/VTween 80/Vnormal saline = 1:1:8), a positive drug group (dexamethasone acetate 10 mg/kg), three isolated blumeatin (4A) groups (including low-, medium-, and high-dose sets), three (±) blumeatin (1) groups, and three (±) sterubin (4) groups. The test compounds were dissolved in a solvent containing 1 mL of ethanol, 1 mL of Tween 80, and 8 mL of normal saline.

All mice were injected intraperitoneally as per group. Five hours later, the animals in each group were smeared with 30 μL of xylene on the inside and outside of the right ear. The left ear was not smeared which served as a control and was given normal saline. One hour after the inflammation was induced, the animals in each group were killed by cervical dislocation, and then the ears were pierced with a puncher of a fixed size. Finally, the tissues cut from the ears were weighed and recorded with a balance, and the degree of swelling and inhibition rate were calculated..

Acknowledgments

This research was supported by the Guizhou University Cultivation Project (grant no. [2020]83) and the National Nature Science Foundation of China (grant no. 32160849).

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.3c01247.

  • HPLC, TLC, 1H NMR, 13C NMR, HMBC, HSQC, IR, and HRMS data of intermediates and target compounds (PDF)

The authors declare no competing financial interest.

Supplementary Material

ao3c01247_si_001.pdf (1.4MB, pdf)

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Supplementary Materials

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