Antioxidant, antibacterial and DNA protective activities of protein extracts from Ganoderma lucidum

Abstract

Crude proteins of cultured mycelia and fruiting bodies of Ganoderma lucidum were investigated for antioxidant, antibacterial and DNA protective activities. It was found that the half maximal inhibitory concentration (IC50) of the mycelia protein and fruiting bodies protein extracts against 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) radical (ABTS(•+)) were 2.47 ± 0.01 and 2.77 ± 0.11 μg protein/ml and against 2,2-diphenylpicrylhydrazyl radical (DPPH(•)) were 2.5 ± 0.01 and 3.42 ± 0.01 μg protein/ml, respectively. The ferric reducing-antioxidant power (FRAP) values of those samples were 1.73 ± 0.01 and 2.62 ± 0.01 μmole trolox/μg protein respectively. Protein hydrolysates prepared by pronase exhibited a weaker antioxidant activity. Both crude proteins showed antibacterial activity, whereas only the mycelia protein extract could protect DNA damage by hydroxyl ((•)OH) radicals. This protein extract was partial purified by Diethyl amino ethyl (DEAE)-Sepharose column and Sulfopropyl (SP)-Sepharose column, obtained major protein with molecular weight about 45 kilo Dalton (kDa). In conclusion, G. lucidum protein extracts have promise potential for applications as antioxidant and antibacterial agents.

Keywords: Antioxidant; DNA protection; Mushroom; Protein purification; Reducing power.

Fig. 1

The antioxidant capacity of crude protein measured as Trolox Equivalents (μmole trolox/μg protein) assay. Each value is expressed as mean ± standard deviation (n = 3). Mean in each sample is significantly different (p < 0.05)

Fig. 2

Twelve percent SDS-PAGE of Ganoderma lucidum protein extracts digested with pronase. a crude mycelia protein extract, lane 1: molecular weight markers; lane 2: pronase 4 μg; lane 3: crude mycelia protein extract; lane 4; 6; 8: control sample at 30 min, 1 h, 2 h; lane 5; 7; 9: digested sample at 30 min, 1 h and 2 h, respectively, b crude fruiting bodies protein extract digested with pronase. Lane 1: molecular weight markers, lane 2: pronase in the reaction, lane 3; 5; 7: control 30 min, 1 h, 2 h, lane 4, 6, 8: treated with pronase at 40 °C for 30 min, 1 and 2 h respectively

Fig. 3

Inhibitory effect of Ganoderma lucidum protein extracts on DNA damage from hydroxyl radicals. a mycelia protein extract, lane 1 plasmid DNA (0.05 μg), lane 2: plasmid DNA exposed Fenton reaction, lane 3: plasmid DNA with Fenton reaction plus 200 μM trolox, lane 4; 5; 6: plasmid DNA with Fenton reaction plus mycelia protein extract 25, 100, 200 μg/ml respectively; b fruiting bodies protein, lane 1: plasmid DNA (0.05 μg), lane 2: plasmid DNA with Fenton reaction, lane 3; 4; 5: plasmid DNA plus fruiting bodies protein 2.5, 25 and 100 μg/ml respectively. OC: open chain; L: linear; SC: supercoil DNA

Fig. 4

DEAE-Sepharose profile of the mycelia protein of Ganoderma lucidum. The proteins were eluted with a linear gradient of 0–1 M NaCl in 10 mM Tris–HCl buffer (pH 7.3). DE1 showed the highest antioxidant activity

Fig. 5

SP-Sepharose profile of DE1 fraction. SP1 and SP2 were eluted with a linear gradient of 0–1 M NaCl in 10 mM ammonium acetate buffer (pH 4.6). SP2 showed the higher antioxidant activity

Fig. 6

Fifteen percent SDS-PAGE of the purified Ganoderma lucidum mycelia protein after SP-Sepharose XL column. Lane M: molecular weight markers; lane 1: crude mycelia protein; lane 2: un-adsorbed protein; lane 3: SP1; lane 4: SP2. Partial purified protein (SP1 and SP2) had major protein with molecular mass about 45 kDa