EPR Laboratory is an integral part of the Center for Physical Chemistry of Biological Systems, located at the Faculty of Physical Chemistry, University of Belgrade. The Center represents the scientific and research team of experts in various aspects of up-to-date problems in the field of physical chemistry of biological systems. The Center also aims to educate young researchers through master, doctoral and post-doctoral programs in the field of biophysical chemistry at the Faculty of Physical Chemistry, University of Belgrade.
Expertise description
- Ex vivo, in vitro and in vivo testing of new anticancer drugs using EPR (electron paramagnetic resonance) and Raman spectroscopy.
- ROS/RNS spin-trapping using EPR.
- EPR detection of ROS/RNS production kinetics.
- EPR spin-labeling of membranes.
- EPR low temperature studies of metalloproteins and protein radicals.
- In vivo EPR spectroscopy of small animals for NO and ROS detection (investigating the pharmacokinetics of nitroxides and detecting NO radicals).
- EPR Oximetry (measuring oxygen ex vivo/in vivo).
- EPR/MRI imaging of trapped radicals.
- EPR measurements of oxidative stress in the brain during cerebral ischemia and reperfusion.
- EPR spectroscopy and imaging – topical applications (directly monitoring the effects of drugs on skin, exploring the effects of UV light on skin, monitoring topical applications of liposomes as delivery systems, monitoring the controlled release from implantable devices – nitroxides as surrogate drugs).
- Physicochemical investigation of biochemical mechanisms of the onset and progression of neurodegenerative diseases (Parkinson’s, Alzheimer’s, multiple sclerosis, amyotrophic lateral sclerosis).
- Physicochemical investigation of morphological and functional changes in the central nervous system in patients with autoimmune disease (myasthenia gravis), psychogenic non-epileptic attacks and Gaucher’s disease.
- Synthesis, functionalization, characterization and ex vivo/in vivo tracking of nanosystems for targeted and controlled drug delivery.
- Creation of a database for different tumors using Raman spectroscopy with the application of advanced computational methods for processing spectra based on neural networks.
- Physicochemical investigation of new potential radioprotectors, as well as the study of differences in the chemical composition of brain structures after irradiation with and without radioprotectors.
- Experimental and theoretical physicochemical testing of pro- and anti-oxidant effects of plant polyphenols, with special emphasis on subclasses of flavonoids and phenolic acids.
- Testing of oscillatory dynamics in a closed reactor and resulting physicochemical detection of short free-radical species.
Skills and competences
- ROS/RNS spin trapping. Method for identification of various types of short-lived free radicals produced in different chemical or biological systems (like homogenates and tissues). Using this approach one can detect which types of radicals are produced in the investigated samples (OH, O2-, NO, SG, H, CH3, CH2OH …) even if several radical types are produced in the same system.
- Detection of ROS/RNS production kinetics by reduction of spin probes. The method in which stable spin probes are used for estimation of the rate of production (or disappearance) of different short-lived radical species. This method is used if we are not certain how many radical types are included in some reaction mechanism and for estimation of the antioxidative activity of different compounds.
- Spin labeling of membranes. The method in which specially designed EPR active molecules called spin labels are used. These molecules are used to label the membrane (plant, animal or liposome) in order to evaluate the membrane properties (e.g. fluidity). This method could reveal the existence of the process of lipid peroxidation by different ROS. The spin labelling method could also be used for labelling proteins and evaluation of their conformational changes.
- Low temperature studies of metalloproteins and protein radicals. Identification of metal-coordination features of metalloproteins that contain transition metal ions (e.g. V, Cr, Mn, Fe, Ni, Cu), metal oxidation states, and types of ligands. Detection and quantification of thiyl and tyrosyl radicals in proteins.
- EPR imaging of small-volume samples (up to 50 µl). These experiments include EPR imaging in the X-band with high sensitivity (nanomolar concentrations). For example, the growth of a spheroid (a model for solid tumours) could be observed using nitroxide spin probes (e.g. how the area of necrosis increases over time).
- In vivo spectroscopy of small animals for detection of NO and ROS. These experiments include investigating the pharmacokinetics of nitroxides as measured using L-band EPR resonators in which small animals (e.g. mice or rats) are placed. It could be observed that the EPR signal diminishes due to the clearance but also due to the reduction by endogenous scavengers, passing through the BBB, etc.
- EPR Oximetry. Simultaneously measuring the production of NO and the pO2 (concentration of oxygen) in sepsis using EPR spectroscopy in the L-band. EPR Oximetry is a technique for measuring the concentration of oxygen in different biological samples (ex vivo and in vivo).
- EPR/MRI imaging of trapped radicals. In vivo imaging of short-lived free radicals is an extremely difficult task. However, until new effective spin traps are found, the solution can be found in using MRI and the paramagnetic properties of trapped radicals as MRI contrast agents. This way, NO in vivo imaging is possible using the combined EPR/MRI approach.
- EPRI of ROS using protected hydroxylamines. Performing EPR measurements of oxidative stress in the brain during cerebral ischemia and reperfusion. The EPR spectrum and EPR image using nitroxide (generated by the oxidation of injected hydroxylamine in the mouse subject) is recorded. Subsequently, the same mouse is recorded on the MRI. Overlapping these EPRI and MRI images, the ischemic and normal side of the brain could be easily observed.
- EPR spectroscopy and imaging – topical applications: (a) Directly monitor the effect of drugs on skin (by detecting drug induced radical formation under pertinent therapeutic conditions); (b) Explore the effect of UV light on skin (UV light presents potent oxidative stress in the skin); (c) Monitor topical applications of liposomes as a delivery system of hydrophilic substances through the skin (nitroxides as surrogate drugs); (d) Early detection of skin malignant melanoma at the initial stage of development; (e) Monitor controlled release from implantable devices – nitroxides as surrogate drugs.
EPR applications