W.W. Sułkowski

Interaction of anticancer drugs with human and bovine serum albumin

Abstract The properties of 5-fluorouracil (5FU), 6-azauracil (6AU), 6-mercaptopurine (6MP), thiouracil (2TU) and thioimidazole (TI), as a quencher of fluorescence of tryptophanyl Trp 214 residue in serum albumin were studied. For the molar ratio [ligand]: [BSA]=30: 1 the decrease of protein fluorescence due to the presence of thiouracil as a quencher attains 90%, while imidazole derivative-thiamazole quenches the BSA fluorescence only by 25%. To estimate the character of the binding between studied quencher and human and bovine serum albumin the Scatchard and Stern–Volmer methods were used. The binding stoichiometry of allopurinol–mercaptopurine–serum albumin complex was studied.

Effect of urea on serum albumin complex with antithyroid drugs: fluorescence study

Abstract Comparative study of the ligand effect on the conformational stability of human (HSA) and bovine (BSA) serum albumin was performed using fluorescence spectroscopy. Tryptophan fluorescence emission spectra of serum albumin in the presence of mercaptopyrimidine (MPI) were recorded at the excitation wavelengths of 280 and 295 nm. The quenching of the fluorescence of protein in the presence of MPI resulted from the interactions with the drug in the subdomain IIA of α-helix where Trp 214 is located. The unfolding process of serum albumin on increasing the concentration of urea (1÷8 M) was studied. It was suggested that the destabilization of serum albumin structure in the presence of increasing urea concentration runs probably through intermediate state at 4÷6 M urea. The binding and quenching constants MPI–HSA and MPI–BSA has been calculated and compared with those obtained under urea-denatured conditions. An unfolding of serum albumin, induced by urea (within the concentration range 1÷5 M), involved diminishing of binding constant by ∼27% for HSA and by ∼49% for BSA. When concentration of urea exceeded 6 M, the binding constant raised by 21 and 27% for HSA and BSA, respectively. Anti-denaturant action of MPI was shown.

Changes of serum albumin affinity for aspirin induced by fatty acid

Saturated fatty acids such as myristic acid play an important role in the pathogenesis of cardiovascular disorders. Using the quenching fluorescence method we examined the influence of myristate on the changes of transporting protein affinity towards aspirin-the most popular anticoagulant. Our results showed that the presence of the myristic acid alters the stability of the anticoagulant-albumin complex. The ranges of [myristate]/[albumin] molar ratio at which the stability of drug-protein complex increases or decreases were determined. The differences in interaction between ligands and human or bovine serum albumins were identified. The competition in binding of ligands with these albumins was also described.

Effective polyelectrolytes synthesised from expanded polystyrene wastes

Chemical modification of used polymer plastics to useful products is a way of effective waste management. To obtain the effective polyelectrolytes the synthesis of sulphonated derivatives of expanded polystyrene wastes was performed. Modification was conducted by means of known methods and products having various contents of sulpho- groups in polymer chain were obtained. Flocculation properties of the anionic type polyelectrolytes obtained were tested. Studies were conducted for coal mine water from the “Kleofas” coal mine and for water from the “Huta Czȩstochowa” plant circulation system. It was found that the polyelectrolytes have good flocculation properties similar to those of anionic type commercial polyelectrolyte. They could be used with good effect for aiding industrial water treatment processes. The optimal concentration of obtained polyelectrolytes was 0.066 mg/dm3. Application of synthesised polyelectrolytes caused a decrease of turbidity and concentration of solved contamination, and improved quality parameter of purified water.

Spectroscopic study of the humification process during sewage sludge treatment

The aim of this work was to study the free radical transition of organic materials during the sewage treatment process. Investigations of sludge from biologic–mechanical sewage treatment plant in Sosnowiec Zagorze were carried out. The course of the humification processes during sewage treatment was studied by electron paramagnetic resonance (EPR) technique. The concentration of free radicals at each process stage and the value g were determined. Sludge samples and extracted fractions of humic acids were examined. Humic acids were extracted from sludge by means of conventional methods elaborated by Stevenson. For study of humic acids structures, besides EPR, the UV–Vis and IR spectroscopy were used.

The spectroscopic studies of humic acid extracted from sediment collected at different seasons

Goczałkowice Reservoir is the biggest water reservoir in the south of Poland. For our studies bottom sediments were collected from eight different places of the reservoir at various seasons of the year. EPR spectroscopy was applied to analyse both quantitatively and qualitatively the humic acids (HA) present in the samples. EPR spectra of the extracted HA exhibited broad lines from the paramagnetic metal ions and narrow lines from free radicals. The values of the free radical concentration obtained for HA amounted to 1.14-13.6 × 1016 spin g(-1) depending on the season and the place of sample collection. The values of the g factor obtained for HA were of the range 2.0027-2.0035. The EPR studies show that HA extracted from bottom sediment collected at various points of the Goczałkowice Reservoir exhibit similar physical-chemical properties. It was also observed that the depth of the reservoir affects the content of the oxygen functional groups as well as the free radical concentration in HA. The identification of the functional groups was done by means of IR. UV/VIS spectroscopy was used to estimate the maturity of the HA.

NMR study of the humification process during sewage sludge treatment

Humic acids, extracted from sludge at the biologic-mechanical sewage treatment plant in Jastrzebie Zdroj, have been investigated by means of (1)H, (13)C and (31)P NMR spectroscopy. Sludge samples for studies were taken from the primary settling tank, the nitrification chamber, the digestion chamber and the sludge drying beds. The (1)H NMR analysis of humic acid extracted from sludge at various stages of sewage treatment confirmed the presence of the functional groups that are characteristic for humic substances, and the analysis showed changes in their relative intensities. The (13)C NMR indicated that the aromatisation of the humic acid increased during sewage treatment. Moreover, the analysis of the (31)P NMR spectra allowed us to observe the changes in the phosphorus groups of the studied humic acids.

A spectroscopic study of phenylbutazone and aspirin bound to serum albumin in rheumatoid diseases.

Interaction of phenylbutazone (PBZ) and aspirin (ASA), two drugs recommended in rheumatoid diseases (RDs), when binding to human (HSA) and bovine (BSA) serum albumins, has been studied by quenching of fluorescence and proton nuclear magnetic resonance ((1)HNMR) techniques. On the basis of spectrofluorescence measurements high affinity binding sites of PBZ and ASA on albumin as well as their interaction within the binding sites were described. A low affinity binding site has been studied by proton nuclear magnetic resonance spectroscopy. Using fluorescence spectroscopy the location of binding site in serum albumin (SA) for PBZ and ASA was found. Association constants K(a) were determined for binary (i.e. PBZ-SA and ASA-SA) and ternary complexes (i.e. PBZ-[ASA]-SA and ASA-[PBZ]-SA). PBZ and ASA change the affinity of each other to the binding site in serum albumin (SA). The presence of ASA causes the increase of association constants K(aI) of PBZ-SA complex. Similarly, PBZ influences K(aI) of ASA-SA complex. This phenomenon shows that the strength of binding and the stability of the complexes increase in the presence of the second drug. The decrease of K(aII) values suggests that the competition between PBZ and ASA in binding to serum albumin in the second class of binding sites occurs. The analysis of (1)HNMR spectral parameters i.e. changes of chemical shifts and relaxation times of the drug indicate that the presence of ASA weakens the interaction of PBZ with albumin. Similarly PBZ weakens the interaction of ASA with albumin. This conclusion points to the necessity of using a monitoring therapy owning to the possible increase of uncontrolled toxic effects.

The influence of dietary habits and pathological conditions on the binding of theophylline to serum albumin

The influence of fatty acids (FA) on theophylline (Th) binding to human serum albumin (HSA) in its high and low affinity binding sites was investigated. The content of studied FA solutions corresponds to the ones associating with different dietary habits and pathological states in vivo. Using fluorescence and (1)H NMR spectroscopy two high and two low affinity binding sites of Th in HSA structure were found. For each site several binding parameters in the absence and presence of FA were estimated. The results showed that the impact of FA on the affinity of HSA towards Th in high affinity binding sites is negligible whereas binding of the drug in low affinity sites decreases significantly in the presence of FA. It was observed that this effect is dependent on the number of fatty acid molecules bound to the protein while the chemical structure of fatty acids contained in the solution plays a minor role.

Effect of temperature on liposome structures studied using EPR spectroscopy

The effect of temperature on liposome structures has been investigated by means of electron paramagnetic resonance spectroscopy with the use of the spin labelling technique. The EPR spectra were recorded on a Bruker EMX spectrometer at the X band in the temperature range 300–340 K. Liposomes were prepared from L?a?phosphatidylcholine dipalmitoyl (1,2?dihexadecanoyl?sn?glycerol?3?phosphocholine) 99% (DPPC), DL?a?phosphatidylcholine dimyristoyl (1,2?ditetradecanoyl?rac?glycerol?3?phosphocholine) 99% (DMPC) and cholesterol (5?cholesten?3s?ol) 99