Two New Monoterpene Glycosides from Qing Shan Lu Shui Tea with Inhibitory Effects on Leukocyte-Type 12-Lipoxygenase Activity

Abstract

We evaluated the inhibitory effect of 12 Chinese teas on leukocyte-type 12-lipoxygenase (LOX) activity. Tea catechins such as epigallocatechin gallate have been known to exhibit leukocyte-type 12-LOX inhibition. Qing Shan Lu Shui, which contains lower catechin levels than the other tested teas, suppressed leukocyte-type 12-LOX activity. To characterize the bioactive components of Qing Shan Lu Shui, leukocyte-type 12-LOX inhibitory activity–guided fractionation of the aqueous ethanol extract of the tea was performed, resulting in the isolation of two new monoterpene glycosides: liguroside A (1) and B (2). The structures of compounds 1 and 2 were characterized as (2E,5E)-7-hydroperoxy-3,7-dimethyl-2,5-octadienyl-O-(α-l-rhamnopyranosyl)-(1″→3′)-(4′″-O-trans-p-coumaroyl)-β-d-glucopyranoside and (2E,5E)-7-hydroperoxy-3,7-dimethyl-2,5-octa-dienyl-O-(α-l-rhamnopyranosyl)-(1″→3′)-(4′″-O-cis-p-coumaroyl)-β-d-glucopyranoside, respectively, based on spectral and chemical evidence. Ligurosides A (1) and B (2) showed inhibitory effects on leukocyte-type 12-LOX activity, with IC50 values of 1.7 and 0.7 μM, respectively.

Involvement of the Hydroperoxy Group in the Irreversible Inhibition of Leukocyte-Type 12-Lipoxygenase by Monoterpene Glycosides Contained in the Qing Shan Lu Shui Tea

“…Proton signals due to an exomethylene (δ 5.19 and 5.18), three vinyls (δ 5.92, 5.63 and 5.59), a methylene (δ 2.32), and three tertiary methyl (δ 1.31, 1.22, 1.22) groups were observed in the aliphatic proton region (Table 1 and Figure S1). The 13 C-NMR resonances of liguroside C were similar to those of liguroside A [7], except for 10 signals of a monoterpene moiety (Table 1 and Figures S2–S5). The monoterpene moiety was assigned as a (5 E )-3,7-dimethyl-1,5-octadiene-3,7-diol analogue, based on the correlations from the HMBC experiment of liguroside C, as shown in Figure 2.…”

“…The monoterpene moiety was assigned as a (5 E )-3,7-dimethyl-1,5-octadiene-3,7-diol analogue, based on the correlations from the HMBC experiment of liguroside C, as shown in Figure 2. Among the signals of monoterpene unit, the signals of the two methyls (δ 24.8, C-8, 9) and those attached to the oxygenated tertiary carbon (δ 81.6, C-7) were close to those due to the methyls (δ 24.1) and the methine (δ 80.6) carbons of liguroside A [7], suggesting that a hydroperoxy group was allocated to the C-7 position of liguroside C; this suggestion was supported by its high-resolution electrospray ionization mass spectroscopy (HRESIMS) analysis. On the basis of these data, the structure of liguroside C was concluded to be (5 E )-7-hydroperoxy-3,7-dimethyl-1,5-octadienyl-3- O -(α- l -rhamnopyranosyl)-(1′′→3′)-(4′′′- O - trans - p -coumaroyl)-β- d -glucopyranoside (Figure 2).…”

“…As shown in Figure 3, liguroside C and liguroside D, dose-dependently inhibited the leukocyte-type 12-lipoxygenase activity with IC 50 values of 2.9 μM and 4.2 μM, respectively. The inhibitory potencies of liguroside C and liguroside D were comparable to those of liguroside A and liguroside B (IC 50 values of 1.7 μM and 0.7 μM) [7]. In our previous study, the leukocyte-type 12-lipoxygenase inhibitory assay-guided fractionation of the 30% aqueous ethanol soluble portion of the ethyl acetate extract was carried out using a reversed-phase HPLC [7].…”

“…The inhibitory potencies of liguroside C and liguroside D were comparable to those of liguroside A and liguroside B (IC 50 values of 1.7 μM and 0.7 μM) [7]. In our previous study, the leukocyte-type 12-lipoxygenase inhibitory assay-guided fractionation of the 30% aqueous ethanol soluble portion of the ethyl acetate extract was carried out using a reversed-phase HPLC [7]. We analyzed the obtained active fraction containing ligurosides A, B, C, and D using a reversed-phase HPLC (Figure S11A).…”

“…A previous report has demonstrated that aqueous extracts of the processed leaves of Ligustrum robustum , dose-dependently scavenged superoxide radicals, inhibited lipid peroxidation, and prevented hemolysis of the red blood cells [2]. We have previously reported that two novel monoterpene glycosides, liguroside A and liguroside B, inhibited the catalytic activity of leukocyte-type 12-lipoxygenase (Figure 1) [7]. Leukocyte-type 12-lipoxygenase not only oxygenates the free fatty acids (such as arachidonic acid) to produce 12-hydroperoxy-5,8,10,14-eicosatetraenoic acid (12-HPETE), but also fatty acids that have esterified to cholesterol in the low-density lipoprotein (LDL) particle, which contributes to the formation of oxidized LDL and the development of atherosclerosis [8,9].…”