Purification of Human Serum Paraoxonase and Effect of Acetylsalicylic Acid on Paraoxonase Activity

Abstract

Paraoxonase was purified from human serum using sepharose-4B-L-tyrosine-1-napthylamine affinity chromatography. This enzyme was purified 1797-fold. SDSpolyacrylamide electrophoresis of the enzyme indicates a single protein staining band with an apparent Mr of 43 kDa. The kinetic properties of the purified enzyme were determined. The V max and K m are 231.5 µ/mg protein (EU) and 3.96 mM using substrate paraoxon respectively. The effect of acetylsalicylic acid on purified human serum paraoxonase has been investigated in vitro. The inhibition and activation effects of acetylsalicylic acid on serum paraoxonase activity were measured spectrophotometrically using paraoxon as the substrate. The phenotypic distribution of paraoxonase was determined using the dual substrate paraoxon and phenyl acetate as substrates. It was observed that acetylsalicylic acid inhibited human serum paraoxonase activity. In addition, an in vivo study was performed for acetylsalicylic acid in Sprague-Dawley rats. It was demonstrated that paraoxonase in the serum of SpragueDawley rat was activated, but activity in liver and heart was significantly inhibited.

Inhibition of paraoxonase 1 by coumarin‐substituted N‐heterocyclic carbene silver(I), ruthenium(II) and palladium(II) complexes

“…For this purpose, some research groups focused on the effects of known compounds on PON1 activity. Aliphatic alcohols, some metal ions, acetylsalicylic acid, coumarin derivatives, anaesthetics, antibiotics, calcium channel blockers, cardiovascular therapeutics, some anticancer agents, some pesticides, some copper complexes, sulfonamide derivatives and antiepileptic drugs have been investigated by various research groups in view of their possible effects on PON1 activity. Additionally, our group investigated the PON1 inhibitory properties of imidazolium salts with coumarin, benzoxazinone or benzotriazole moieties and we showed that coumarin‐substituted imidazolium salts are stronger inhibitors of PON1 than corresponding 4‐chloromethylene coumarins .…”

The Differentially Effect of Some Antilipid Drugs on Activity of the Novel Synthesized Paraoxonase (PON1)-Inorganic Hybrid Nanoflowers

“…to separate serum. The obtained serum was used to purify PON1 enzyme via the three-phase separation method [27,28].…”

“…The reactions were incubated for 15 minutes at 37 o C and the absorbance changes were recorded at 412 nm and enzymes activities were determined. The % Activity-Inhibitor concentration graphs were plotted versus three drugs using the obtained data and IC50 values were calculated (Table 4) [27,28,38].…”

Synthesis and evaluation of novel 2-[(1,2,4-triazol-3-yl)thio]acetamide derivatives as potential serum paraoxonase-1 (PON1) activators

“…Activity measurement is based on establishing the changes in absorption of formed p-nitrophenol as a result of the reaction of PON1 with paraoxon at 412 nm. p-Nitrophenol the molar extinction coefficient is e = 18.290M −1 cm −1 and an enzyme unit is number of micromole of hydrolysis paraoxon in per minute [41][42][43].…”

“…Human paraoxonase 1 was eluted with the solution 25 mM Tris/HCl (pH 8.0)/10 mM CaCl 2 at 0.5mL/min. Fractions (3 mL each) were collected as 3 mL, and those with the highest PON1 activity were combined [42,43].…”