Chromatographic Separation of Breynia retusa (Dennst.) Alston Bark, Fruit and Leaf Constituents from Bioactive Extracts

Abstract

Breynia retusa (Dennst.) Alston (also known as Cup Saucer plant) is a food plant with wide applications in traditional medicine, particularly in Ayurveda. Extracts obtained with four solvents (dichloromethane, methanol, ethyl acetate and water), from three plant parts, (fruit, leaf and bark) were obtained. Extracts were tested for total phenolic, flavonoid content and antioxidant activities using a battery of assays including 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), ferric reducing antioxidant power (FRAP), cupric reducing antioxidant capacity (CUPRAC), total antioxidant capacity (TAC) (phosphomolybdenum) and metal chelating. Enzyme inhibitory effects were investigated using acetylcholinesterase (AChE), butyrylcholinesterase (BChE), tyrosinase, α-amylase and α-glucosidase as target enzymes. Results showed that the methanolic bark extract exhibited significant radical scavenging activity (DPPH: 202.09 ± 0.15; ABTS: 490.12 ± 0.18 mg Trolox equivalent (TE)/g), reducing potential (FRAP: 325.86 ± 4.36: CUPRAC: 661.82 ± 0.40 mg TE/g) and possessed the highest TAC (3.33 ± 0.13 mmol TE/g). The methanolic extracts were subjected to LC-DAD-MSn and NMR analysis. A two-column LC method was developed to separate constituents, allowing to identify and quantify forty-four and fifteen constituents in bark and fruits, respectively. Main compound in bark was epicatechin-3-O-sulphate and isolation of compound was performed to confirm its identity. Bark extract contained catechins, procyanidins, gallic acid derivatives and the sulfur containing spiroketal named breynins. Aerial parts mostly contained flavonoid glycosides. Considering the bioassays, the methanolic bark extract resulted a potent tyrosinase (152.79 ± 0.27 mg kojic acid equivalent/g), α-amylase (0.99 ± 0.01 mmol acarbose equivalent ACAE/g) and α-glucosidase (2.16 ± 0.01 mmol ACAE/g) inhibitor. In conclusion, methanol is able to extract the efficiently the phytoconstituents of B. retusa and the bark is the most valuable source of compounds.

LC-ESI-QTOF-MS/MS Analysis, Cytotoxic, Antiviral, Antioxidant, and Enzyme Inhibitory Properties of Four Extracts of Geranium pyrenaicum Burm. f.: A Good Gift from the Natural Treasure

“…Moreover, Geranium pyrenaicum ethyl acetate extract was more toxic to cancer cells than Geranium robertianum ethyl acetate extract, and the difference was especially marked in case of HeLa cells, 32.34 vs. 217 µg/mL, respectively. Geranium robertianum aqueous extracts (infusion, decoction) showed variable cytotoxicity against cancer cells (GI 50 45.68-380 µg/mL) [24], whereas, we have observed consistent anticancer activity of Geranium pyrenaicum aqueous extract towards all tested cell lines with high selectivity (Table 5). In another paper based on similar study design, Graça, Dias, Barros, Calhelha, Santos and Ferreira [23] reported the cytotoxicity of Geranium molle L. extracts towards the same cancer panel and PLP2 cells.…”

“…The acetone extract exerted highest cytotoxicity against all tumor (GI 50 -50% of growth inhibition, 57-71 µg/mL) and non-tumor (GI 50 176 µg/mL) cell lines [24]. Interestingly, cytotoxicity of Geranium robertianum n-hexane extract was noticeably lower towards cancer cells (GI 50 151-179 µg/mL) [24] than we have observed (Table 5) for Geranium pyrenaicum nhexane extract (CC 50 53.72-75.46 µg/mL), in case of HeLa cells, this difference was vs. 73.28 µg/mL, respectively. Moreover, Geranium pyrenaicum ethyl acetate extract was more toxic to cancer cells than Geranium robertianum ethyl acetate extract, and the difference was especially marked in case of HeLa cells, 32.34 vs. 217 µg/mL, respectively.…”

“…The highest selectivity towards all cancer cell lines (SI 4.5-10.8) was observed for Geranium pyrenaicum aqueous extract with the FaDu cells being the most sensitive (CC 50 40.22 µg/mL). The Geranium maculatum ethanolic extract was reported to possess moderate (IG 50 (50% inhibitory growth) 60.2 µg/mL) on MDA-MB-231 (human breast cancer) cells [68], which is similar to the activity we have observed for Geranium pyrenaicum methanol extract on FaDu cells (CC 50 66.92 µg/mL). Graça, Barros, Calhelha, Dias, Carvalho, Santos-Buelga, Santos and Ferreira [24] reported the cytotoxicity of organic (n-hexane, dichloromethane, ethyl acetate, acetone and methanolic) and aqueous (infusion and decoction) extracts from Geranium robertianum L. towards MCF-7 (breast adenocarcinoma), NCI-H460 (nonsmall cell lung cancer), HeLa and HepG2 (hepatocellular carcinoma) cancer cell lines, as well as hepatotoxicity against non-tumor porcine liver primary cells (PLP2).…”

“…With the increasing polarity of solvent used for extraction an increase of SI could be observed. In case of Geranium pyrenaicum methanolic extract (GP-ME) a significant anticancer activity on all cell lines could be observed (SI 3.6-7.2) and the highest activity (CC 50 66.92 µg/mL) and selectivity found on FaDu cells. The highest selectivity towards all cancer cell lines (SI 4.5-10.8) was observed for Geranium pyrenaicum aqueous extract with the FaDu cells being the most sensitive (CC 50 40.22 µg/mL).…”

“…In case of Geranium pyrenaicum methanolic extract (GP-ME) a significant anticancer activity on all cell lines could be observed (SI 3.6-7.2) and the highest activity (CC 50 66.92 µg/mL) and selectivity found on FaDu cells. The highest selectivity towards all cancer cell lines (SI 4.5-10.8) was observed for Geranium pyrenaicum aqueous extract with the FaDu cells being the most sensitive (CC 50 40.22 µg/mL). The Geranium maculatum ethanolic extract was reported to possess moderate (IG 50 (50% inhibitory growth) 60.2 µg/mL) on MDA-MB-231 (human breast cancer) cells [68], which is similar to the activity we have observed for Geranium pyrenaicum methanol extract on FaDu cells (CC 50 66.92 µg/mL).…”