Purification and characterization of dihydropyrimidine dehydrogenase enzyme from sheep liver and determination of the effects of some anaesthetic and antidepressant drugs on the enzyme activity

Figures & data

Figure 1. (A) Lineweaver–Burk graph of five different uracil concentrations and in constant NADPH concentration. (B) Lineweaver–Burk graph of five different NADPH concentrations and in constant uracil concentration.

Figure 2. SDS-PAGE bands of DPD enzyme from sheep liver. Gel stained using the silver staining procedure. Lane 1: affinity chromatograph; lane 2: standard proteins (SDS-PAGE: sodium dodecyl sulphate polyacrylamide gel electrophoresis, DPD: dihydropyrimidine dehydrogenase enzyme).

Table 1. Summary of purification steps of dihydropyrimidine dehydrogenase (DPD) from sheep liver.

Purification steps	Volume (mL)	Enzyme activity (EU/mL)	Protein (mg/mL)	Total activity (EU)	Total protein (mg)	Specific activity (EU/mg)	Purification fold	Yield (%)
Crude homogenate	27.0	0.073	42.00	1.971	1134.0	1.73 × 10^−3	1.0	100
Ammonium sulphate precipitation	14.0	0.055	29.65	0.770	415.1	1.85 × 10^−3	1.07	39.1
DEAE cellulose ion exchange chromatography	4.5	0.028	0.078	0.126	0.351	0.359	207.5	6.4
2′,5′-ADP-sepharose-4B affinity chromatography	1.0	0.012	0.017	0.012	0.017	0.705	407.5	6.1

Figure 3. Standard R_f-Log MW graph for the determination of molecular weight of sheep liver DPD.

Figure 4. (A) The effect of pH on the activity of sheep liver DPD. (B) The effect of ionic strength on the activity of sheep liver DPD. (C) The effect of temperature on the activity of sheep liver DPD. (D) Determination of stable pH for sheep liver DPD.

Table 2. The half maximal inhibitory concentration (IC₅₀) of some antidepressant and anaesthetic drugs against dihydropyrimidine dehydrogenase (DPD) from sheep liver.

Drugs	IC50 values (μM)
Escitalopram	1736.11
Fluoxetine	13.24
Mirtazapine	86.65
Haloperidol	99.03
Propofol	0.21
Lidocaine	15.07