Antibiotics Used in Patients after Surgery and Effects of Human Serum Paraoxonase-I (PON1) Enzyme Activity.

Abstract

Paraoxonase (PON; arilesterase, [EC 3.1.8.1]) is an enzyme from the group arilesterases (ARE). This enzyme is capable of hydrolyzing paraoxone which is the active metabolite of parathion, an organic phosphorus insecticide. PON activity was found to be low in individuals prone to development of atherosclerosis such as diabetes, familial hypercholesterolemia and kidney disorders. It was noted that PON enzyme activity decreases in relation to age increase in adults. PON enzyme activity is approximately half of that in newborns and premature babies. Approximately one year after birth, it reaches the adult level. It can be said that PON1 has significant role on living organisms. For this reason, many studies on interactions of PON-drugs are needed. In this article, our aim is to investigate in vitro effects of four pharmaceutically active agents (fosfomycin, cefuroxime axetil, cefaclor monohydrate, and cefixime) which are often used in patients after surgery on human serum paraoxanase-I (PON1) enzyme activity. In this article, we purify paraoxonase-I enzyme from human serum by using ammonium sulfate precipitation (in the range of 60-80%), ion exchange and gel filtration chromatography. We use electrophoresis to check the purity of the enzyme. We investigate the paraoxonase activity of the enzyme at 412 nm the inhibition effects of the active substances. Paraoxone is used as the substrate. Activity measurements arw made at different inhibitor concentrations related to inhibitor studies and % Activity- [I] graphs are drawn for drug active substances. Lineweaver-Burk graphics are used to determine the Ki constants. Finally, to determine the types of inhibition we interpret these graphs. The active agents used after surgery decreased the PON1 enzyme activity. They showed different inhibition mechanism. The inhibition mechanism of fosfomycin and cefaclor monohydrate was noncompetitive, cefixime was uncompetitive and cefuroxime axetil was a competitive inhibitor. The IC50 values for fosfomycin, cefuroxime axetil, cefaclor monohydrate, and cefixime were calculated to be 31.5 mM, 1.03 mM, 4.18 mM and 0.781 mM, respectively, and the Ki constants were determined to be 27.98 ± 12.25 mM, 2.20 ± 0.22 mM, 4.81 ± 2.25 mM and 1.12 ± 0.32 mM, respectively. The IC50 and Ki values showed that cefixime active agent has the maximum inhibition. In this study, we have detected that cefuroxime axetil inhibited competitively in vitro paraoxonase activity of this enzyme. According to this information, we thought that cefuroxime axetil linked to the active site of the enzyme. Fosfomycin and cefaclor monohydrate can be attached with amino acids out of the active site of the enzyme because they inhibit enzyme noncompetitively. Cefixime can be attached only to the enzyme-substrate complex because it inhibits enzyme uncompetitively.