Chemical composition, antioxidant potential, macromolecule damage and neuroprotective activity of Convolvulus pluricaulis

Abstract

Herbal medicines are known to mitigate radical induced cell damage. Hence identification and scientific validation of herbal medicines contribute to better use in Ayurvedic/Unani research. In the present study, we investigated antioxidant and anti-apoptotic properties of Convolvulus pluricaulis (C. pluricaulis). C. pluricaulis exhibited antioxidant potential evident by free radical scavenging activities. C. pluricaulis pretreatment inhibited H₂O₂ induced macromolecule damage such as plasmid DNA damage and AAPH induced oxidation of bovine serum albumin and lipid peroxidation of rat hepatic tissues. Further to identify the neuroprotective properties of C. pluricaulis, SHSY5Y cells were treated with H₂O₂ with or without pretreatment of C. pluricaulis. The C. pluricaulis pretreatment at 50 μg/ml dose exhibited 50% cell survival against 100 μM H₂O₂ challenge for 24 h and it also decreased the lactate dehydrogenase leakage. Further C. pluricaulis pretreatment restored and regulated the antioxidant and apoptosis markers such as SOD, CAT, p53, and caspase-3 and inhibited, reactive oxygen species generation and depolarization of the mitochondrial membrane. C. pluricaulis possess a high content of flavonoids and polyphenols and GC-MS and FTIR analysis showed a wide variety of compounds which may contribute to the observed effects.

Keywords: Antioxidant; Convolvulus pluricaulis; FTIR; GC-MS; Neuroprotection; SH-SY5Y.

Graphical abstract

Fig. 1

GC–MS chromatogram of Convolvulus pluricaulis 70% ethanolic extract.

Fig. 2

FTIR spectrum of Convolvulus pluricaulis 70% ethanolic extract.

Fig. 3

Protective effect of C. pluricaulis against AAPH induced pUC19 DNA damage.

Fig. 4

Protective effect of C. pluricaulis against H₂O₂ induced Protein oxidation.

Fig. 5

a Cytotoxic effects H₂O₂ on SHSY5Ycells.b Dose dependent protective effect of treatment with CP on H₂O₂ induced cytotoxicity in SHSY5Y cells, the cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. c Protective effect of C. pluricaulis pretreatment on H₂O₂ induced cytotoxicity by LDH leakage assay. The data are represented as mean ± SD of three independent experiments. ^#P < 0.01 versus control group,*P < 0.01 versus 100 μM H₂O₂ treated group. d Effects of CP pretreatment and 100 μM H₂O₂ induced morphological alterations in SH-SY5Y neurons observed by phase contrast microscopy.

Fig. 6

Pre-treatment of C. pluricaulis for the restoration of SOD and catalase enzyme activities in SH-SY5Y cells challenged with 100 μM H₂O₂. The data are represented as mean ± SD of three independent experiments. ^#P < 0.01 versus control group, *P < 0.01 versus 100 μM H₂O₂ treated group.

Fig. 7

Estimation of intracellular ROS production using DCFH2DA in SHSY5Y cells pre-treated with C. pluricaulis on 100 μM H₂O₂ challenge by spectrofluorimeter. The ROS generation was monitored by fluorescent microscope. The data are represented as mean ± SD of three independent experiments. ^#P < 0.01 versus control group,*P < 0.01 versus 100 μM H₂O₂ treated group.

Fig. 8

Estimation of mitochondrial membrane potential in SHSY5Y cells with pre-treatment of C. pluricaulis on 100 μM H₂O₂. The fluorescence intensity was determined using a spectrofluorimeter. The membrane potential was monitored by fluorescent microscope. The data are represented as mean ± SD of three independent experiments. ^#P < 0.01 versus control group,*P < 0.01 versus 100 μM H₂O₂ treated group.

Fig. 9

Protective effect of C. pluricaulis on DNA damage induced by 100 μM H₂O₂ in SHSY5Y cells. (a) Control cells without any treatment (b) Cells with 100 μM H₂O₂ treatment (c) Cells were pre-treated with C. pluricaulis for 2 h at 50 μg/assay and treated with 100 μM H₂O₂ (duration: 24 h) Tail length (50 μm) Bars. ^#P < 0.05 versus control group, *P < 0.05 versus H₂O₂ treated group.

Fig. 10

The protective effect of pre-treatment of C. pluricaulis on H₂O₂ induced expression of oxidative stress marker proteins SOD, CAT and apoptosis marker proteins caspase 3, p53 analyzed by Western blotting. (b,c,d,e) The band intensity is calculated by the Image-J software. The data are represented as mean ± SD of three independent experiments. ^#P < 0.05 versus control group, *P < 0.05 versus H₂O₂ treated group.

Fig. 11

Neuroprotective effects of Convolvulus pluricaulis are mediated via its anti-oxidant and anti-apoptotic activity against H₂O₂ induced neurotoxicity in SH-SY5Y human neuronal cells.