**4. PPAR-γ ligand-binding activity of compounds 1–39 isolated from** *G. glabra*

H-5′ (δH 6.72) and C-1′ (δC 121.0)/C-3′ (δC 138.7)/C-4′ (δC 149.0), H-6′ (δH 7.25) and C-β (δC 138.4)/ C-2′ (δC 148.7), and methoxy protons and C-2′. Therefore, the structure of **1** was assigned as

Compound **5** was isolated as a yellow amorphous powder with a molecular formula of

of an isoflavan skeleton at δH 4.43 (ddd, *J* = 10.2, 3.4, 2.2 Hz, H-2a), 4.07 (dd, *J* = 10.2, 10.2 Hz, H-2b), 3.54 (m, H-3), 3.07 (dd, *J* = 15.5, 11.1 Hz, H-4a), and 2.90 (ddd, *J* = 15.5, 5.0, 2.2 Hz, H-4b). In addition, the spectrum of **5** indicated signals that we assigned to two aromatic protons at δ<sup>H</sup> 7.58 and 6.49 (each s), *ortho*-coupled aromatic protons at δH 6.87 and 6.32 (each d, *J* = 8.2 Hz), and a 2,2-dimethylpyran ring at δH 6.63 and 5.65 (each 1H, d, *J* = 9.8 Hz) and δH 1.40 and 1.38

myl group, which was attached at C-5′, as determined by the HMBC correlations between the formyl proton signal and C-4′ (δC 163.6)/C-5′ (δC 115.4)/C-6′ (δC 133.9) (**Figure 5**). The circular dichroism (CD) profile of **5** was the same as that of synthetic 5′-formylglabridin prepared by formylation of **20**, indicating that the absolute configuration at C-3 was *R*. Therefore, the structure of **5** was assigned as 5′-formyl glabridin. It was notable that **5** was the first naturally

H and 13C NMR signals at δH 9.77 and δC 195.3 indicated the presence of a for-

(each 3H, s). Compound **5** and glabridin (**20**) have similar features in their 1

occurring isoflavan with a formyl group in the B-ring portion of the compound.

Compound **7** was isolated as a yellow powder with a molecular formula of C20H20O<sup>5</sup>

mined by HRESIMS. Compound **7** had an absorption maxima at 313 and 276 nm in its UV

H NMR spectrum of **5** indicated signals characteristic

H NMR spectra.

deter-

3,3′,4,4′-tetrahydroxy-2′-methoxy-5-prenylchalcone.

**Figure 5.** Key HMBC correlations of **1**, **5**, **7**, and **8** [1].

82 Biological Activities and Action Mechanisms of Licorice Ingredients

determined by HRESIMS. The 1

C21H20O<sup>5</sup>

Moreover, the 1

Compounds **5**, **7**, **11**, **18**, **19**, **26**, **28**, **31**–**33**, **36**, and **37** showed significant PPAR-γ ligand-binding activity. Among these compounds, the prenylflavone derivative licoflavanone A (**31**) was the most potent (**Figure 6**). These active compounds likely contributed the most to the PPAR-γ ligandbinding activity of the EtOH extract. The isoflavone derivative, kanzonol X (**19**), and flavanone derivative, glabrol (**32**), both had two prenyl units and exhibited potent ligand-binding activity. Hispaglabridin B (**24**) and xambioona (**35**), in which two prenyl units were cyclized to form two six-membered rings, exhibited weaker ligand-binding activities than **19** and **32** did, suggesting

**Figure 6.** PPAR-γ ligand-binding activity of compounds **1–39** at 2 ( ) and 10 ( ) μg/mL with a GAL-4-PPAR-γ chimera assay [1]. Troglitazone (TRG) at 0.5, 1.0, and 2.0 μM was used as a positive control, and dimethyl sulfoxide at 1 mL/L was used as a solvent control. Values are means ± SD, *n* = 4 experiments. Statistical significance is indicated by \* (*p* < 0.05) or \*\* (*p* < 0.01) as determined by Dunnett's multiple comparison test.

that the two non-cyclic prenyl moieties were necessary for the potent activity of these compounds (**Figure 7**). Taking together all the above data, the PPAR-γ ligand-binding activity of the phenolic compounds was affected by slight differences in the substitution groups on the aromatic rings.

**Figure 7.** PPAR-γ ligand-binding activity of **19**, **24**, **34**, and **35** isolated from *G. glabra* roots [1]. Values in parentheses are the relative luminescence intensities at 10 μg/mL.
