Bartosz Urbaniak

Analysis of the factors that significantly influence the stability of fluoroquinolone–metal complexes

The aim of the present study was to evaluate factors contributing to the differences between the overall stability constants (logbeta(pqr)) of the fluoroquinolone-metal ion complexes. The experiments were performed using potentiometric titration method in wide pH range. The overall stability constants (logbeta(pqr)) were determined using the Hyperquad program. Complexation equilibria of eight different fluoroquinolones with six divalent and trivalent metal ions were investigated in this study. The authors employed a multifactorial ANOVA analysis, fixed effect model to describe the influence of particular variables affecting the stability of the analyzed complex species. Four different variables were set at different levels labeled. The ligand number (LF) was the first factor. LF determined the number of fluorochinolone molecules in the complex structure, and could take the values 1, 2 or 3. The second factor (Me) was connected with the type of the metal ion bonded in the complex. Since six different metal cations were studied, the Me factor was described with six levels. The number of hydrogen or hydroxide groups substituted into the complex molecule was the third variable (HR) with many levels labeled: q, a, s, d, f and g. The last factor FQ described the type of the fluorochinolone used for complex formations. All variables analyzed here were statistically significant (p value lower than 0.01), which indicates that all of them strongly affect the logbeta(pqr) value. Binary interactions (LF-Me, LF-FQ, Me-HR and Me-FQ) between variables were also stated, which suggests that the effects of these variables were higher than we could calculate based on the effect of each variable alone. The ANOVA analysis has shown that the following factors Me, LF and HR were the most important for the stability of the fluoroquinolone-metal ion complexes. It was also found that according to the FQ factor (type of ligand molecule) all analyzed fluoroquinolones formed stable complexes with metals. It was proved that the application of ANOVA for the entire complexation profile of analyzed fluoroquinolones with polyvalent metal ions was a valid technique for detecting the statistically significant differences in the complexation profiles. Such information may be very useful for better understanding and interpretation of differences in bioavailability of fluoroquinolones and their interactions with antacids and other multimineral drugs.

Activation of Prodrug Treosulfan at pH 7.4 and 37°C Accompanied by Hydrolysis of Its Active Epoxides: Kinetic Studies with Clinical Relevance

Treosulfan (TREO), originally registered for treatment of ovarian cancer, is currently being investigated for conditioning prior to hematopoietic stem cell transplantation. TREO is a prodrug, which undergoes a pH- and temperature-dependent two-step conversion to active monoepoxide [S,S-EBDM, (2S,3S)-1,2-epoxybutane-3,4-diol-4-methanesulfonate] and diepoxide [S,S-DEB, (2S,3S)-1,2:3,4-diepoxybutane]. In this paper, the kinetics of the nonenzymatic transformation of TREO at pH 7.4 and 37°C were studied for the first time including the effects of the TREO concentration, buffer concentration, ionic strength, and the presence of NaCl. Transformation of TREO was well described by a kinetic model, which included first-order reactions for TREO activation, that is, TREO → S,S-EBDM → S,S-DEB, and pseudo-first-order reactions for the hydrolytic decomposition of S,S-EBDM and S,S-DEB. In contrast to the two-step activation of TREO, the hydrolysis of epoxides was influenced by electrolytes. In phosphate-buffered saline, decomposition of S,S-EBDM and S,S-DEB (mean half-lives 25.7 and 15.4 h) proceeded much slower than their formation (mean half-lives 1.5 and 3.5 h). In conclusion, the kinetics of the nonenzymatic transformation of TREO in the presence of plasma electrolytes cannot contribute to the very low levels of S,S-EBDM and S,S-DEB observed in patient plasma. The results also indicate that elimination of TREO proceeds primarily via conversion to S,S-EBDM.

A new method for determination of hyaluronidase activity in biological samples using capillary zone electrophoresis

The aim of the study was to develop a new capillary zone electrophoresis (CZE) method for determination of enzymatic activity of hyaluronidase. The method permits monitoring of the process of hyaluronic acid digestion by hyaluronidase. Studies were performed using CZE instrument equipped with capillary of 64.5 cm total length, 56 cm effective length and internal diameter 75 µm. Separation was performed in the phosphate buffer (pH 8.10) in the electric field of 20 kV, λ = 220 nm. The procedure was based on mixing a known quantity of hyaluronic acid and an aliquot of hyaluronidase solution, followed by obtaining CZE profiles after a known period of incubation (0.5 h). The activity of hyaluronidase was calculated using multiple regression analysis in which sizes of the peaks of the main degradation products were used. The newly developed method was fully validated and it is appropriate to evaluate the activity of hyaluronidase originating from different sources with high precision and accuracy. t-Tests showed that there were no significant differences between results obtained using turbidimetric, viscosimetric and the new CZE method. The developed method is characterized by a short duration of analysis, low volume of analyzed sample, small amount of buffers used and low cost of analysis.

Comparison of the pharmacokinetics of paracetamol from two generic products in patients after total gastric resection.

Gastrectomy leads to pathophysiological changes within the alimentary tract, which may affect drug absorption and pharmacokinetic parameters (PK). The need to apply orally administered analgesics in this group of patients is often related with alternative application of currently available generic products. Thus, from the clinical point of view it is necessary to evaluate the PK of these drugs to confirm their equivalence. The aim of the study was therefore an analysis of the pharmacokinetics of paracetamol from two generic products in patients after total gastric resection. The research was carried out on two groups of patients after gastrectomy with Roux-en-Y reconstruction (n = 30; mean [SD] age, 63.0 [11.5] years; weight, 67.6 [13.7] kg; and height, 166.4 [9.1] cm). The patients received paracetamol in a single orally administered dose of 1,000 mg. Blood samples were collected within 6 h of drug administration. The concentration of paracetamol and paracetamol glucuronide in the blood plasma was marked by means of a validated high-pressure liquid chromatography with ultraviolet detection (HPLC-UV). The main PK for paracetamol in group 1 (n = 17) and 2 (n = 13) were as follows: C(max), 9.46 (3.66) and 12.79 (5.32) μg/ml, respectively (p = 0.0517); AUMC(0-t), 77.64 (30.37) and 51.01 (15.76) μg h(2)/ml (p = 0.0046); AUC(0-inf), 41.61 (23.52) and 30.28 (9.74) μg h/ml (p = 0.0862); t(max), 1.68 (0.63) and 0.50 (0.25) h (p < 0001). The obtained C(max) and AUC values in patients after gastrectomy were reduced in comparison with healthy subjects. Total gastrectomy therefore affected the pharmacokinetics of paracetamol administered in tablets. In our patients, we also observed significant differences between the PK of paracetamol and two generic preparations. These two drugs can thus be used interchangeably, but with caution.

The pharmacokinetics of the effervescent vs. conventional tramadol/paracetamol fixed-dose combination tablet in patients after total gastric resection

BACKGROUND Tramadol/paracetamol is a fixed-dose combination prescribed for the relief of moderate to severe pain. The combination acts synergistically and guarantees the rapid onset of paracetamol and the prolonged analgesic effect of tramadol with good tolerability. These drugs are often used in various formulations in the treatment of patients with postoperative pain, e.g. after stomach resection. Gastrectomy leads to pathophysiological changes within the alimentary tract, which may affect the process of drug absorption. The aim of the research was an analysis of the pharmacokinetics of tramadol/paracetamol from effervescent and conventional tablets in patients after total gastrectomy. METHODS The research was carried out on patients after gastrectomy with Roux-en-Y reconstruction. The patients received two tramadol/paracetamol fixed-dose combination tablets in a single orally administered dose of 75/650 mg (2 × 37.5/325 mg). The patients were subjected to one of the two study drug group with: I. effervescent tablet (ET) (n = 14; mean [SD] age, 63.4 [10.1] years; weight, 75.5 [15.3]kg; and BMI, 26.0 [4.6]kg/m(2)) and II. conventional tablet (CT) (n = 12; mean [SD] age, 66.8 [7.7] years; weight, 79.8 [17.8]kg; and BMI, 27.4 [5.3]kg/m(2)). Blood samples were collected within 10 h after the drug administration. The plasma concentrations of tramadol and paracetamol were measured with validated HPLC (high-performance liquid chromatography) method with UV detection. RESULTS The comparison of the paracetamol and tramadol C(max) ratio for the ET group with that of the CT group gave ratios of 1.16 [90% confidence interval (CI) 1.06, 1.27] and 0.86 (90% CI 0.72, 1.02), respectively. The comparison of the paracetamol and tramadol AUC(0-t) ratio for the ET group with that of the CT group showed ratios of 0.99 (90% CI 0.88, 1.10) and 1.00 (90% CI 0.82, 1.22), respectively. The comparison of the difference for the effervescent and conventional formulation gave an estimated decrease in t(max) of 0.5 h for paracetamol and 0.13 h for tramadol. CONCLUSIONS In view of the changes in the pharmacokinetics of paracetamol and tramadol in the patients after gastric resection for both formulations compared the conventional tablet seems to be more appropriate due to the comparable rate of absorption of both substances, higher concentrations of tramadol and comparable exposure to paracetamol.

The feature selection approach for evaluation of potential rheumatoid arthritis markers using MALDI-TOF datasets

BACKGROUND The selection of the most representative mass profiles, in rheumatoid arthritis (RA) serum samples was developed. This allows for selection and identification of potential biomarkers in RA serum samples. METHODS The RA and controls samples were analyzed using MALDI-TOF. Two different protein elution procedures utilizing ZipTips (E1 and E2) were examined. The statistical evaluation of data was performed using different feature selection (FS) methods in combination with different classifiers, while identification of selected masses was performed using MALDI-TOF-TOF. RESULTS Utilization of proposed statistical strategy allowed for the selection of different masses according to FS method and elution procedure. Obtained masses were further subjected for targeted identification. The panel of proteins were identified as potential markers. The role of these proteins was discussed in relation to pathomechanism of RA. CONCLUSION Application of advanced biostatistical analysis of obtained MALDI-TOF datasets, resulted with targeted selection of potential RA biomarkers. Five proteins were identified due the E1 procedure, and six proteins were identified due the E2 procedure, respectively. The panel of identified proteins suggest that presented statistical methodology and proteomic strategy was correct and gave valid results. Obtained results may contribute to development of clinically useful multicomponent diagnostic tool.

Serum free amino acid levels in rheumatoid arthritis according to therapy and physical disability

Background: In presented study the amino acid analysis was performed in serum derived from rheumatoid arthritis patients (RA) according to undertaken therapy and classification of physical disability. The results were compared with previously published data. Methods: The levels of 31 free amino acids were determined in 50 serum samples derived from RA subjects and 51 controls. The RA patients were divided into two groups according to the therapy (methotrexate/leflunomide, infliximab/adalimumab/etanercept/tocilizumab, prednisolone/NSAID) and classification of physical disability of the patients. Levels of amino acids were measured by LC‐MS/MS. The obtained results were subjected to multivariate statistical tests. Results: According to the therapy that was being used, threonine differentiated RA patients treated with methotrexate/leflunomide ‐ infliximab/adalimumab/etanercept/tocilizumab (p = 0.00954) and infliximab/adalimumab/etanercept/tocilizumab ‐ prednisolone/NSAID (p = 0.03109), while tryptophan differentiated RA patients treated with methotrexate/leflunomide ‐ infliximab/adalimumab/etanercept/tocilizumab (p = 0.01723). In the functional classification, arginine differentiated RA samples between class III and IV (p = 0.02332), while glycine differentiated them between class I+II and III of the Steinbrocker functional classification (p = 0.03366). Conclusions: An analysis of the metabolome profile requires the use of validated bioanalytical methods that are strictly dedicated for this purpose. The obtained results are not accidental (p value less than 0.05), and all of the selected amino acids play an important role in inflammation and immune response. It is suggested that studied amino acids can be considered as a markers for diagnosis of RA and monitoring pharmacotherapy of the disease.

Amino Acids in Cerebrospinal Fluid of Patients with Aneurysmal Subarachnoid Haemorrhage: An Observational Study

Background The authors are aware of only one article investigating amino acid concentrations in cerebrospinal fluid (CSF) in patients with ruptured intracranial aneurysms, and this was published 31 years ago. Since then, both management of subarachnoid haemorrhage (SAH) and amino acid assay techniques have seen radical alterations, yet the pathophysiology of SAH remains unclear. Objective To analyse the pattern of concentrations of amino acids and related compounds in patients with different outcomes following aneurysmal SAH. Methods 49 CSF samples were collected from 23 patients on days 0–3, 5, and 10 post-SAH. Concentrations of 33 amino acids and related compounds were assayed by liquid chromatography tandem mass spectrometry in patients with good [Glasgow Outcome Scale (GOS) 1–3] and poor (GOS 4–5) outcome. Results Of the 33 compounds assayed, only hydroxyproline and 3-aminoisobutyric acid appeared not to increase significantly following SAH. In poor outcome patients, we found significantly higher concentrations of aspartic acid (p = 0.038), glutamic acid (p = 0.038), and seven other compounds on days 0–3 post-SAH; glutamic acid (p = 0.041) on day 5 post-SAH, and 2-aminoadipic acid (p = 0.033) on day 10 post-SAH. The most significant correlation with GOS at 3 months was found for aminoadipic acid on day 10 post-SAH (cc = −0.81). Conclusion Aneurysmal rupture leads to a generalised increase of amino acids and related compounds in CSF. The patterns differ between good and poor outcome cases. Increased excitatory amino acids are strongly indicative of poor outcome.