Castanopsis lamontii Water Extract Shows Potential in Suppressing Pathogens, Lipopolysaccharide-Induced Inflammation and Oxidative Stress-Induced Cell Injury

Abstract

Castanopsis lamontii is traditionally used to prevent inflammatory diseases such as periodontitis and pharyngitis by residents in southwest China. However, little scientific evidence has been found to support this. In this research, the antibacterial activities of Castanopsis lamontii water extract (CLE) were assessed using the micro-dilution method. The anti-inflammatory and antioxidant activities of CLE were investigated in RAW264.7 cells. Key bioactive compounds in CLE were also explored. Results showed that CLE was capable of inhibiting the periodontitis pathogen Porphyromonas gingivalis and the pharyngitis pathogen β-hemolytic Streptococcus. It suppressed lipopolysaccharide-induced inflammation in RAW 264.7 cells via inactivating the TLR4/NF-κB pathway. Besides, it reduced oxidative stress-induced cell injury via scavenging reactive oxygen species. Chemical composition analysis revealed that CLE was rich in epicatechin and procyanidin B2. Further studies confirmed that epicatechin predominantly contributed to the antibacterial activities of CLE, while procyanidin B2 was mainly responsible for the anti-inflammatory activities of CLE. Both compounds contributed to the antioxidant activities of CLE. Acute oral toxicity tests proved that CLE was practically non-toxic. These results provide experimental evidences of the health-beneficial effects of CLE and may help promote the application of CLE in the food and health industries.

Keywords: Castanopsis lamontii; anti-bacteria; anti-inflammation; antioxidant; epicatechin; procyanidin B2.

Figure 1

The buds of Castanopsis lamontii (A) and its infusion (B).

Figure 2

HPLC-UV and ultra-performance liquid chromatography-quadrupole-time-of-flight/mass spectrometry (UPLC-Q-TOF/MS) analysis of the Castanopsis lamontii water extract (CLE). (A) HPLC chromatograms of gallic acid, caffeine and catechins; (B) HPLC chromatograms of procyanidin B2 (PB2); (C) Total ion chromatogram of CLE (lower), ion chromatogram of CLE at m/z 289.071 (middle) and ion chromatogram of CLE at m/z 577.134 (upper) at ESI⁻; (D) Total ion chromatogram of the PB2 standard (lower) and ion chromatogram of the PB2 standard at m/z 577.134 (upper); (E) ESI⁻-MS spectra of peaks at retention time 2.718 min for PB2 (lower), 2.589 min for C (middle), and 2.946 min for EC (upper), respectively; (F) ESI⁻-MS/MS spectra of peaks at retention time 2.768 min for PB2 (lower), 2.611 min for C (middle), and 2.996 min for EC (upper), respectively; (G) ESI⁻-MS/MS spectra of the peak at retention time 2.718 min for the PB2 standard (lower) and ESI⁻-MS spectra of the peak at retention time 2.711 min for the PB2 standard (upper).

Figure 3

Anti-inflammatory activities of CLE, EC, and PB2. (A–C) The effects on nitric oxide (NO), prostaglandin E2 (PGE2), and tumor necrosis factor α (TNF-α) secretion of LPS-pretreated or co-treated RAW 264.7 cells; (D,E) The impacts on the expression of toll-like receptor 4 (TLR4), p-NF-κB (p65), NF-κB (p65), inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) served as the loading control. The same letter within each column indicates no significant difference (p > 0.05). Comp. is short for compounds. 4CLE (2CLE) is short for 400 (200) μg/mL CLE. The dosages of EC and PB2 were identical amount of EC and PB2 in 400 μg/mL CLE, which were 120 μg/mL and 34.4 μg/mL, respectively.

Figure 4

Antioxidant activities of CLE, EC, and PB2. (A) The effects on the lactate dehydrogenase (LDH) release of 0.5 mM H₂O₂-pretreated or co-treated RAW 264.7 cells; (B) The effects on the intracellular reactive oxygen species (ROS) levels of 0.5 mM H₂O₂-pretreated or co-treated RAW 264.7 cells; (C) The effects on the viability of 1 mM H₂O₂-pretreated or co-treated RAW 264.7 cells; (D–F) The hydrogen peroxide and free radical scavenging activities of CLE, EC, and PB2. The same letter within each column indicates no significant difference (p > 0.05). 4CLE (2CLE) is short for 400 (200) μg/mL CLE. The dosages of EC and PB2 were identical amount of EC and PB2 in 400 μg/mL CLE, which were 120 μg/mL and 34.4 μg/mL, respectively.