Papers by Malgorzata Jaworska
Cytokine, Feb 1, 2022
OBJECTIVE Paroxysmal nocturnal hemoglobinuria (PNH) is a clonal non-malignant disease in which he... more OBJECTIVE Paroxysmal nocturnal hemoglobinuria (PNH) is a clonal non-malignant disease in which hematopoietic cell apoptosis may play an important pathophysiological role. Previous studies of the content of phosphatidylinositol (3,4,5)-trisphosphate (PI(3,4,5)P3) indicated the possibility of remote transmission of anti-apoptotic signals between pathological and normal hematopoietic progenitors. METHODS The study determined the plasma levels of beta chemokines and cytokines in N = 19 patients with PNH and 31 healthy controls. The research material was peripheral blood plasma (EDTA) stored at -80 °C until the test. Beta chemokine and cytokine concentrations were tested in duplicate with Bio-Plex Pro Human Cytokine Assay (Bio-Rad, Hercules, CA, USA) using a Luminex 200 flow cytometer and xPONENT software (Luminex Corporation, Austin, TX, USA). In peripheral blood CD34+ cells we tested the proportions of PI(3,4,5)P3+ and Annexin binding apoptotic phenotype using FC and phosflow. RESULTS Compared to the control group, the PNH group showed a significant increase in the plasma concentration of some beta chemokines and cytokines, including MIP-1alpha/CCL3, eotaxin/CCL11, MCP1/CCL2, IL4 and G-CSF. In the group of PNH patients, a significant decrease in the concentration of some cytokines was also observed: RANTES/CCL5, MIP-1beta/CCL4, PDGF-BB and IL9. At the same time, the plasma concentrations of the chemokine IP-10/CXCL10 and the cytokines IFN-gamma, TNF, IL6 and IL10 showed no significant deviations from the values for the control group. Anti-apoptotic phenotype and phosphatidylinositol (3,4,5)-trisphosphate content in PNH clone of CD34+ cells were associated with the level of CCL3 and negatively associated with CCL5, CCL4, PDGF-BB and IL9. CONCLUSIONS This data suggest the existence of apoptotic and PI(3,4,5)P3 imbalance in PNH CD34+ cells driven by anti-apoptotic cytokine biosignature in PNH. Plasma cytokines and intracellular enzymes that regulate the phosphoinositide pathways may become a therapeutic target in PNH.
Journal of Chromatography A, Aug 1, 1999
Capillary electrophoresis has been applied to separate and determine N-acetylcysteine (NAC) and r... more Capillary electrophoresis has been applied to separate and determine N-acetylcysteine (NAC) and related impurities. Determination conditions were found to be optimum with 100 mmol / l borate as the buffer, pH 8.40. The limit of detection was established for each substance examined. The method has been validated by examining linearity ranges, precision and repeatability. The method was used to determine the content of NAC in, and purity of, pharmaceutical preparations. The major impurities (N,N-diacetylcystine, N,S-diacetylcysteine and cystine) were determined at levels of 0.1%.
PubMed, 1993
A reversed-phase high-performance liquid chromatography method for determination of benzodiazepin... more A reversed-phase high-performance liquid chromatography method for determination of benzodiazepine derivatives has been developed. Oxazepam and diazepam was determined in serum and oxazepam in urine.
Acta Haematologica Polonica, 2015
Acta Chromatographica, 2011
Summary Because of the problems with stability and solubility, some of the amino acids (AAs) used... more Summary Because of the problems with stability and solubility, some of the amino acids (AAs) used in parenteral nutrition products are often replaced by their acetylated forms. These include N-acetyltyrosine (NAT) and N-acetylcysteine (NAC). This leads to a need to develop new analytical methods for rapid and easy determination of these substances in the presence of common AAs. Capillary electrophoresis (CE) is one of the techniques that have been successfully applied to the assay of multi-component samples containing AAs and their derivatives. This paper discusses a new CE method for the simultaneous determination of two acetylated AAs in solutions for parenteral nutrition. A background electrolyte (BGE) was developed based on borate buffer with alkaline pH. The method is selective and enables the separation and assay of analytes without special sample pretreatment. Validation parameters confirmed sufficient precision and accuracy of the method. Its applicability was verified by testing several medicinal...
Annals of agricultural and environmental medicine : AAEM, 2013
Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it p... more Determination of the plasma amino acid (AA) levels in Huntington's disease (HD) can make it possible to find the metabolic markers used in early diagnosis. The aim of the presented study was to determine the AA profile in plasma samples from HD patients and presymptomatic carriers, compared to healthy subjects. The AA profile was analyzed with HPLC. The study concerned 59 participants: 30 subjects with abnormal CAG repeats expansion (>36) in the HTT gene, and 29 healthy subjects. Each participant was analyzed with regard to the parameters characterizing the metabolic state and protein metabolism, such as: urea, creatinine, glucose, total protein, TSH (thyroid-stimulating hormone), cortisol, ESR (erythrocyte sedimentation rate), and CRP (C-reactive protein). Simple statistical comparisons showed 5 AA to be significantly lower in the HD group, compared to the control group, i.e.: Asn, His, Leu, Ser, Thr. Creatinine and creatinine clirens were found to be lower in the HD group, ...
Molecules
The method of using high-performance liquid chromatography with a charged aerosol detector method... more The method of using high-performance liquid chromatography with a charged aerosol detector method (HPLC-CAD) was developed for the separation and determination of phospholipids isolated from cell membranes. The established cell lines—normal and neoplastic prostate cells and normal skin fibroblasts and melanoma cells—were selected for the study. Chromatographic separation was performed in the diol stationary phase using a gradient elution based on a mixture of n-hexane, isopropanol and water with the addition of triethylamine and acetic acid as buffer additives. Taking the elements of the Folch and Bligh–Dyer methods, an improved procedure for lipid isolation from biological material was devised. Ultrasound-assisted extraction included three extraction steps and changed the composition of the extraction solvent, which led to higher recovery of the tested phospholipids. This method was validated by assessing the analytical range, precision, intermediate precision and accuracy. The ana...
International Journal of Applied Ceramic Technology
Acta poloniae pharmaceutica, 2012
High-performance liquid chromatography (HPLC) is a technique most frequently used for the assessm... more High-performance liquid chromatography (HPLC) is a technique most frequently used for the assessment of biologically active peptides. Owing to the ionic character of these compounds, they may also be separated and assayed using capillary electrophoresis (CE), which offers very high efficiency, short analysis time and low consumption of reagents, and is used increasingly more often. The paper describes the combination of HPLC and CE in order to increase the efficiency of the separation of complex mixture of peptides (active substance and its related impurities). The developed two-dimensional HPLC-CE technique was employed for the analysis of the impurities of octreotide, a cyclic octapeptide used in therapy. Because distinct separation mechanisms are used, the two-dimensional technique ensures higher separation efficiency and a more comprehensive impurity profile of the medicinal product than either of the techniques used separately.
Acta poloniae pharmaceutica
Currently, glutamine is thought to be conditionally indispensable amino acid. However, the use of... more Currently, glutamine is thought to be conditionally indispensable amino acid. However, the use of glutamine in clinical practice encounters serious problems related to its limited solubility and instability in water solutions. Therefore, glutamine-containing dipeptides, which have better solubility and stability in the solution, have been introduced into the medical practice. High concentration of L-alanyl-L-glutamine in the medicinal product and thermal sterilization during the manufacturing process are the reasons of many impurities formation--the products of deamidation, racemization, cyclization etc.. The purpose of the study was to develop a single step CE method that could be applied in the purity control of L-alanyl-L-glutamine containing parenteral infusions instead of three routinely used LC methods. After systematic optimization separation conditions were selected: 100 mM borate buffer at pH 8.5 with 0.4 mM diaminopropane as EOF modifier. The method developed allows the se...
Advances in Clinical and Experimental Medicine
Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiam... more Background. Oxidative stress accompanies neurodegeneration and also causes abnormalities in thiamine-dependent processes. These processes have been reported to be diminished in the brains of patients with several neurodegenerative diseases. Objectives. The aim of this work was to conduct a comparative analysis of the impact of supplemented thiamine on the viability of human B lymphocytes with CAG abnormal expanded huntingtin gene (mHTT) (GM13509) and control, B lymphocytes without mHTT (GM14467) through the following studies: determination of the supplemented thiamine concentrations, which are effective for cell growth stimulation after incubation in thiamine deficit conditions; determination of cell capability to intake the exogenous thiamine; evaluation of exogenous thiamine influence on the profile of the genes related to thiamine and energy metabolism; determination of ATP synthesis and activities of thiamine-dependent enzymes, KGDHC and BCKDHC in the intact cells and upon the exogenous thiamine. Material and methods. The following methods were used: EZ4U test for cell growth analysis; HPLC for determination of thiamine intake and ATP synthesis, qRT-PCR for evaluation of the gene profiles and spectrophotometric method for KGDHC and BCKDHC activities determination. Results. Maximal cell growth stimulation was observed at 2.5 mM in GM14467 up to 135% of the control culture and at 5.0 mM in GM13509 cells up to 165% of the control culture. Native levels of total ATP and KGDHC and BCKDHC activities in both cell types were comparable and did not changed upon thiamine deficit or supplementation. GM13509 cells showed more of an increase in growth stimulation upon thiamine supplementation than GM14467 cells and this effect was reflected in the increase of intracellular thiamine concentration. Conclusions. The above results and reported changes in expression of GAPDH, IDH1 and SLC19A3 genes observed upon thiamine deficit conditions suggest that intracellular thiamine status and energy metabolism can have a role in HD pathogenesis.
Journal of Pharmaceutical and Biomedical Analysis, Aug 1, 2009
Sodium caprylate and N-acetyltryptophan are the most frequently used stabilizers that protect the... more Sodium caprylate and N-acetyltryptophan are the most frequently used stabilizers that protect the albumin from aggregation or heat induced denaturation. In turn citrates - excipients remaining after fractionation process - can be treated as by-product favoring leaching aluminum out of glass containers whilst albumin solution is stored. With ionic nature these substances have all the markings of a subject for capillary electrophoresis analysis. Thus CE methods were proposed as new approach for quality control of human albumin solution in terms of determination of stabilizers and citrates residue. Human albumin solutions both 5% and 20% from various manufacturers were tested. Indirect detection mode was set to provide sufficient detectability of analytes lacking of chromophores. As being anions analytes were separated with reversed electroosmotic flow. As a result of method optimization two background electrolytes based on p-hydroxybenzoic acid and 2,6-pyridinedicarboxylic acid were selected for stabilizers and citrates separation, respectively. The optimized methods were successfully validated. For citrates that require quantification below 100microM the method demonstrated the precision less than 4% and the limit of detection at 4microM. In order to check the new methods accuracy and applicability the samples were additionally tested with selected reference methods. The proposed methods allow reliable quantification of stabilizers and citrates in human albumin solution that was confirmed by method validation as well as result comparison with reference methods. The CE methods are considered to be suitable for quality control yet simplifying and reducing cost of analysis.
Talanta, Dec 1, 2010
The paper proposes a new method for amino acid determination which can be applied for amino acid ... more The paper proposes a new method for amino acid determination which can be applied for amino acid profiling in solutions for parenteral nutrition. The MEKC method based on a mixed micellar system was developed for the separation of 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatized amino acids. Background electrolyte was based on tris-borate buffer with high alkaline pH. Sodium dodecyl sulfate micelles were modified using 1,2-hexanediol as a co-surfactant. The effect of the modifier on amino acid migration was studied with respect to hydrophobicity of the analytes. The modifier appeared to be suitable to improve the separation of AQC-tagged amino acids without an adverse effect on buffer ionic strength or EOF velocity. The method was successfully validated and applied for amino acid profiling in medicinal preparations for parenteral nutrition. The results obtained were compared with a reference chromatographic method (amino acid analyser).
Acta poloniae pharmaceutica
High-performance liquid chromatography (HPLC) is a technique most frequently used for the assessm... more High-performance liquid chromatography (HPLC) is a technique most frequently used for the assessment of biologically active peptides. Owing to the ionic character of these compounds, they may also be separated and assayed using capillary electrophoresis (CE), which offers very high efficiency, short analysis time and low consumption of reagents, and is used increasingly more often. The paper describes the combination of HPLC and CE in order to increase the efficiency of the separation of complex mixture of peptides (active substance and its related impurities). The developed two-dimensional HPLC-CE technique was employed for the analysis of the impurities of octreotide, a cyclic octapeptide used in therapy. Because distinct separation mechanisms are used, the two-dimensional technique ensures higher separation efficiency and a more comprehensive impurity profile of the medicinal product than either of the techniques used separately.
Acta biochimica Polonica, 2014
Regimen-related mucosal toxicity is extremely common following cytotoxic chemotherapy and radioth... more Regimen-related mucosal toxicity is extremely common following cytotoxic chemotherapy and radiotherapy. Mucositis is as an important determinant of the inflammatory response and infectious complications in cancer treated patients. Most assessment scales for mucosal damage are focussed on oral mucositis, since it is easy to evaluate. Measuring gastrointestinal musocal damage objectively remains difficult because it cannot be seen directly or readily detected. One of potential non-invasive biomarkers of gastrointestinal mucosal damage is plasma citrulline level. Citrulline is an amino acid produced by small bowel enterocytes. Low concentration of free circulating citrulline signifies severe intestinal mucosal damage in humans with nonmalignant disorders, such as villous atrophy-associated diseases, short bowel syndrome, Crohn's disease, and is used in follow-up after small bowel transplantation. The plasma citrulline level is a reliable and objective biochemical marker of enterocy...
Talanta, 2010
The paper proposes a new method for amino acid determination which can be applied for amino acid ... more The paper proposes a new method for amino acid determination which can be applied for amino acid profiling in solutions for parenteral nutrition. The MEKC method based on a mixed micellar system was developed for the separation of 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) derivatized amino acids. Background electrolyte was based on tris-borate buffer with high alkaline pH. Sodium dodecyl sulfate micelles were modified using 1,2-hexanediol as a co-surfactant. The effect of the modifier on amino acid migration was studied with respect to hydrophobicity of the analytes. The modifier appeared to be suitable to improve the separation of AQC-tagged amino acids without an adverse effect on buffer ionic strength or EOF velocity. The method was successfully validated and applied for amino acid profiling in medicinal preparations for parenteral nutrition. The results obtained were compared with a reference chromatographic method (amino acid analyser).
Journal of Separation Science, 2005
Preservatives are used to protect pharmaceutical formulations from microbial attack during the pe... more Preservatives are used to protect pharmaceutical formulations from microbial attack during the period of administration to the patient. Because of their biological activity, preservatives have to be identified and assayed according to the same rules as apply to active components. A number of methods for separation of preservatives are reported, to account for the heterogeneity of their chemical structures. A capillary electrophoretic method was devised for simple and simultaneous qualification and quantification of the preservatives most often included in pharmaceuticals, such as benzyl alcohol, parabens, phenol, m-cresol, chlorobutanol, thimerosal. After systematic method development, the electrophoretic conditions were defined as: 50 mM borate buffer pH 9.0 containing 20 mM SDS. Separations were performed at a temperature of 20 degrees C and with detection at 214 nm. Preservatives under examination can be analyzed within a 10 min run. The method was successfully validated and applied to the determination of preservatives in a number of pharmaceuticals. Results from the CE method were compared with those from reference methods.
Journal of Pharmaceutical and Biomedical Analysis, 2009
Sodium caprylate and N-acetyltryptophan are the most frequently used stabilizers that protect the... more Sodium caprylate and N-acetyltryptophan are the most frequently used stabilizers that protect the albumin from aggregation or heat induced denaturation. In turn citrates - excipients remaining after fractionation process - can be treated as by-product favoring leaching aluminum out of glass containers whilst albumin solution is stored. With ionic nature these substances have all the markings of a subject for capillary electrophoresis analysis. Thus CE methods were proposed as new approach for quality control of human albumin solution in terms of determination of stabilizers and citrates residue. Human albumin solutions both 5% and 20% from various manufacturers were tested. Indirect detection mode was set to provide sufficient detectability of analytes lacking of chromophores. As being anions analytes were separated with reversed electroosmotic flow. As a result of method optimization two background electrolytes based on p-hydroxybenzoic acid and 2,6-pyridinedicarboxylic acid were selected for stabilizers and citrates separation, respectively. The optimized methods were successfully validated. For citrates that require quantification below 100microM the method demonstrated the precision less than 4% and the limit of detection at 4microM. In order to check the new methods accuracy and applicability the samples were additionally tested with selected reference methods. The proposed methods allow reliable quantification of stabilizers and citrates in human albumin solution that was confirmed by method validation as well as result comparison with reference methods. The CE methods are considered to be suitable for quality control yet simplifying and reducing cost of analysis.
Journal of Chromatography A, 2003
Ketosteril is an enteral medicinal product indicated for prevention and therapy in chronic renal ... more Ketosteril is an enteral medicinal product indicated for prevention and therapy in chronic renal insufficiency in connection with a low protein diet. Tablets of Ketosteril contain five essential amino acids like: Lys, His, Thr, Trp, Tyr and another five amino acids in the form of their hydroxy and keto analogues as calcium salts, that are: alpha-ketoleucine, alpha-ketoisoleucine, alpha-ketovaline, alpha-ketophenylalanine and alpha-hydroxymethionine. The composition of Ketosteril tablets is routinely tested with three LC methods. Capillary electrophoretic method seems to be a good alternative for amino acids and their analogues determination in multicomponent pharmaceuticals because of short analysis time and the possibility to assay all components during a single run without any pretreatment. Electrophoresis was performed in 50 microm I.D. fused-silica capillaries with 65 cm distance to the detector. Capillaries were installed in Waters Quanta 4000 electrophoretic equipment with a positive power supply and on-line UV detection at 214 nm. Separations were done in a buffer containing 40 mM Tris and 160 mM boric acid titrated with NaOH to pH 10. The method developed allows the separation of all investigated analytes with an efficiency of n = 230,000 and 20 min analysis time. The method was applied for determination of all components of Ketosteril in commercial tablets.
Journal of Chromatography A, 2005
The paper presents a rapid method for the determination of commonly used synthetic food dyes by m... more The paper presents a rapid method for the determination of commonly used synthetic food dyes by micellar electrokinetic capillary chromatography. Detection and separation conditions allowing complete resolution of 15 synthetic food colourants were investigated. The effect of different surfactants on the analytes mobility in relation to their structure was tested. After optimization procedure a dual micellar system was selected. All food dyes were separated in less then 20 min using a fused silica capillary in the borate/dodecylsulfate/deoxycholate buffer containing acetonitrile as organic modifier. The detection wavelength was set at 210 nm. The method was successfully validated by determination of linearity ranges, detection limits, precision and repeatability for all colourants tested. In order to apply the method for pharmaceutical analysis a sample pretreatment procedures were found. Liquid pharmaceuticals were used as it or just after dilution with water. From tablets or capsules the colourants were isolated by adsorption on acidic aluminium oxide. The method was used for identification and if possible for quantification the synthetic food dyes in pharmaceuticals. The analytes are detectable at a concentration level 0.3-0.8 g ml −1 .
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Papers by Malgorzata Jaworska