The processes of lipid peroxidation in the cells of Chlorobium limicola IMV K-8 under the influence of copper (II) sulphate

Abstract

The effect of stressors, including heavy metal ions such as Cu2+, promotes activation of free radical processes in the cells of microorganisms, which causes changes in their physiological and biochemical properties and the structure of bacterial membranes. The aim of this work was to assess the influence of copper (II) sulphate on intensity of lipid peroxidation (LPO) of Chlorobium limicola IMV K-8 by measuring the content of primary (conjugated dienes and lipid hydroperoxides) and secondary lipid peroxidation products (TBA-reactive products). Microorganisms were cultivated at a temperature of 28 °C in GSB cultivation medium with exposure to light of wavelength 700–800 nm and intensity 40 lux. A suspension of C. limicola ІМV К-8 cells in the phase of exponential growth was treated for one hour with metal salt solution in concentrations 0.05–0.50 mM for investigation of the influence of copper (II) sulphate on its physiological and biochemical properties. The control samples did not contain any copper (II) sulphate. Biomass was determined by turbidity of diluted cell suspension by application of photoelectric colorimeter KFK-3. A mixture of n-heptane and isopropyl alcohol was added into cell-free extract for conjugated dienes determination. The samples were incubated at room temperature and centrifuged. Water was added into the supernatant and the samples were stirred. Ethanol was added to the heptanes phase and adsorption was measured at 233 nm. The content of lipid hydroperoxides was determined by a method based on protein precipitation by trichloroacetic acid followed by addition of ammonium thiocyanate. The concentration of TBA-reactive products in the cell-free extracts was determined by color reaction with malondialdehyde and thiobarbituric acid exposed to high temperature and acidity of the medium, which causes formation of trimetinic adduct with maximal absorption at 532 nm. It was shown that when CuSO4 was added to the incubation medium the content of conjugated dienes and lipid hydroperoxides increased with the enhancement of salt concentration. However, its value decreased by the seventh and eighth days of cultivation. The content of TBA-reactive products under the influence of copper (II) sulphate varied depending on the duration of cultivation and concentration of the metal. Its highest quantity was observed on the eighth day of cultivation. Thus it was determined that under the influence of CuSO4 the content of conjugated dienes, lipid hydroperoxides and TBA-reactive products increases. This indicates the increased activity of lipid peroxidation processes and the free radical chain reaction damage mechanism to lipids under these conditions.