Phenolic Compounds in Organic and Aqueous Extracts from Acacia farnesiana Pods Analyzed by ULPS-ESI-Q-oa/TOF-MS. In Vitro Antioxidant Activity and Anti-Inflammatory Response in CD-1 Mice

Delgadillo Puga Claudia, Cuchillo-Hilario Mario, Navarro Ocaña Arturo, Medina-Campos Omar Noel, Nieto Camacho Antonio, Ramírez Apan Teresa, López-Tecpoyotl Zenón Gerardo, Díaz Martínez Margarita, Álvarez-Izazaga Marsela Alejandra, Cruz Martínez Yessica Rosalina, Sánchez-Quezada Vanessa, Gómez Francisco Enrique, Torre-Villalvazo Iván, Furuzawa Carballeda Janette, Camacho-Corona María Del Rayo, Pedraza-Chaverri José, Delgadillo Puga Claudia, Cuchillo-Hilario Mario, Navarro Ocaña Arturo, Medina-Campos Omar Noel, Nieto Camacho Antonio, Ramírez Apan Teresa, López-Tecpoyotl Zenón Gerardo, Díaz Martínez Margarita, Álvarez-Izazaga Marsela Alejandra, Cruz Martínez Yessica Rosalina, Sánchez-Quezada Vanessa, Gómez Francisco Enrique, Torre-Villalvazo Iván, Furuzawa Carballeda Janette, Camacho-Corona María Del Rayo, Pedraza-Chaverri José

Abstract

Background: Acacia farnesiana (AF) pods have been traditionally used to treat dyspepsia, diarrhea and topically for dermal inflammation. Main objectives: (1) investigate the antioxidant activity and protection against oxidative-induced damage of six extracts from AF pods and (2) their capacity to curb the inflammation process as well as to down-regulate the pro-inflammatory mediators.

Methods: Five organic extracts (chloroformic, hexanic, ketonic, methanolic, methanolic:aqueous and one aqueous extract) were obtained and analyzed by UPLC-ESI-Q-oa/TOF-MS. Antioxidant activity (DPPH•, ORAC and FRAP assays) and lipid peroxidation (TBARS assay) were performed. Assessment of anti-inflammatory properties was made by the ear edema induced model in CD-1 mice and MPO activity assay. Likewise, histological analysis, IL-1β, IL-6, IL-10, TNF-α, COX measurements plus nitrite and immunohistochemistry analysis were carried out.

Results: Methyl gallate, gallic acid, galloyl glucose isomer 1, galloyl glucose isomer 2, galloyl glucose isomer 3, digalloyl glucose isomer 1, digalloyl glucose isomer 2, digalloyl glucose isomer 3, digalloyl glucose isomer 4, hydroxytyrosol acetate, quinic acid, and caffeoylmalic acid were identified. Both organic and aqueous extracts displayed antioxidant activity. All extracts exhibited a positive effect on the interleukins, COX and immunohistochemistry assays.

Conclusion: All AF pod extracts can be effective as antioxidant and topical anti-inflammatory agents.

Keywords: Acacia farnesiana pods; antioxidant and anti-inflammatory activities; bioactive compounds; polyphenols.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Scavenging capacity of the DPPH• radical (A–G) and inhibitory concentration 50% (IC50) values (H) of different extracts from Acacia farnesiana pods. The data are the mean with the standard deviation of three independent repetitions. CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract; Q = quercetin. a,b,c,d,e Different letters showed significant difference (p < 0.05) in the same extract at different concentrations (Kruskal–Wallis test).
Figure 1
Figure 1
Scavenging capacity of the DPPH• radical (A–G) and inhibitory concentration 50% (IC50) values (H) of different extracts from Acacia farnesiana pods. The data are the mean with the standard deviation of three independent repetitions. CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract; Q = quercetin. a,b,c,d,e Different letters showed significant difference (p < 0.05) in the same extract at different concentrations (Kruskal–Wallis test).
Figure 2
Figure 2
Total antioxidant activity of the extracts from Acacia farnesiana pods by oxygen radical absorbance capacity (ORAC) and ferric-reducing antioxidant power (FRAP) assays. Q = quercetin; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. a,b,c,d,e,f Different letters showed significant difference (p < 0.05) in the same extract at different concentrations (Kruskal–Wallis test).
Figure 3
Figure 3
Representative images of reactive oxygen species (ROS) produced on porcine kidney cells exposed to H2O2 and to 200 ppm of extracts of Acacia farnesiana pods (A) and ROS quantification using 50, 100 and 200 ppm of extracts (B). H2O2 = hydrogen peroxide; Q = quercetin; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract; AE = aqueous extract. a,b,c,d Different letters showed significant difference (p < 0.05) among extracts and queretin at the same concentration (Kruskal–Wallis test).
Figure 3
Figure 3
Representative images of reactive oxygen species (ROS) produced on porcine kidney cells exposed to H2O2 and to 200 ppm of extracts of Acacia farnesiana pods (A) and ROS quantification using 50, 100 and 200 ppm of extracts (B). H2O2 = hydrogen peroxide; Q = quercetin; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract; AE = aqueous extract. a,b,c,d Different letters showed significant difference (p < 0.05) among extracts and queretin at the same concentration (Kruskal–Wallis test).
Figure 4
Figure 4
Inhibition of lipid peroxidation (A–G) and inhibitory concentration 50% (IC50) values (H) of the extracts from AF pods and standard quercetin, quantified by lipid peroxidation (TBARS) production. CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. Q = quercetin. The data are the mean with the standard error of three independent repetitions. * p < 0.05 and ** p < 0.01 vs. control group.
Figure 5
Figure 5
Effect of different extracts from Acacia farnesiana pods and the anti-inflammatory indomethacin (1 mg) on TPA-induced ear edema model. (A) ear edema measured at 4 h after TPA treatment and (B) oxidative enzyme myeloperoxidase (MPO) activity in supernatants of homogenates from TPA-treated ears. Levels of (C) interleukin-1β (D), interleukin-6 (E), interleukin-10 and (F) TNF-α in supernatants of homogenates from ears after treatment with different extracts from AF pods. TPA = 12-O-tetradecanoylphorbol acetate; In = indomethacin; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. Each bar represents the mean ± standard deviation (n = 6). * p < 0.05 and ** p < 0.001 with respect to TPA group.
Figure 6
Figure 6
Histological features of mice ear tissue (A) stained with hematoxylin-eosin (200× magnification) on mice ear tissue thickness by the effect of different extracts of Acacia farnesiana pods on the TPA-induced ear edema model (B). TPA = 12-O-tetradecanoylphorbol acetate; In = indomethacin; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. Each bar represents the mean ± standard deviation (n = 6). ** p < 0.001 with respect to TPA group.
Figure 7
Figure 7
TNF-α expression (golden yellow color) in the mice ear cells due to the effect of different extracts of Acacia farnesiana (AF) pods (3 mg/ear) on the TPA-induced ear edema model (100× magnification). CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract.
Figure 8
Figure 8
Effect of different Acacia farnesiana extracts on prostaglandin production. Cx = Celecoxib; CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. The data are the mean with the standard deviation of three independent repetitions. Statistical difference was calculated in relation to COX at 30 and 100 ppm. * p < 0.05. ** p < 0.001.
Figure 9
Figure 9
(A) Viability of RAW 264.7 cells treated with of lipopolysaccharides (LPS) of E. coli (1 µg/mL), oleanolic acid or different extracts (25 µg/mL) from Acacia farnesiana pods. (B) Nitrite concentration in supernatants of RAW 264.7 cells treated with LPS of E. coli (1 µg/mL), oleanolic acid (Ole) different extracts (25 µg/mL) from Acacia farnesiana (AF) pods. CE = chloroformic extract; HE = hexanic extract; KE = ketonic extract; ME = methanolic extract; MEAE = methanolic:aqueous extract and AE = aqueous extract. Each bar represents mean ± standard deviation (n = 6). * p < 0.05 between extracts respect to LPS.

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