In vitro effects of some drugs on human erythrocyte glutathione reductase

Figures & data

Figure 1.  SDS-PAGE photograph: Lane 1, purified enzyme from Sephadex G-200 gel filtration; Lane 2, standard proteins: Bovine carbonic anhydrase (29 kDa), chicken ovalbumin (45 kDa), bovine albumin (66 kDa).

Figure 2.  Activity% versus drug concentration regression analysis graphs for glutathione reductase (GR) in the presence of different drug concentrations.

Figure 3.  Lineweaver–Burk graphs using three different drugs (A: ketotifen; B: meloxicam; C: phenyramidol–HCl; D: gadopentetic acid) concentrations for determination of K_i.

Table 1.  Purification scheme of glutathione reductase (GR) from human erythrocyte.

Purification step	Total volume (mL)	Activity (U/mL)	Total activity (U)	Protein (mg/mL)	Specific activity (U/mg)	Yield (%)	Purification factor
Haemolysate	86	0.053	4.56	35.6	0.0015	100	1
Ammonium sulphate precipitation (30–70%)	37	0.064	2.368	28.9	0.002	52	1.46
2′,5′-ADP Sepharose 4B affinity chromatography	6	0.255	1.53	0.535	0.047	33.5	31.5
Sephadex G-200 gel filtration chromatography	4	0.211	0.844	0.0521	0.314	18.5	209

Table 2.  K_i values obtained from Lineweaver–Burk graphs for glutathione reductase (GR) in the presence of three fixed inhibitors and five substrate concentrations for different drugs.

Drugs	IC50 (mM)	Ki (mM)	Inhibition type
Ketotifen	0.012	0.0008 ± 0.00021	Non-competitive
Meloxicam	0.029	0.0061 ± 0.00127	Non-competitive
Phenyramidol–HCl	0.99	0.4340 ± 0.0890	Competitive
Gadopentetic acid	138	28.84 ± 4.6900	Non-competitive