A high-throughput miniaturized liquid-liquid extraction procedure followed by a simple ultra-high performance liquid chromatography method coupled with fluorescence detection for bioanalytical analysis of all tocopherol isomers and retinol in human serum has been developed and validated. In the extraction procedure, a synthetic internal standard tocol was used, which does not occur in the human body. The separation of structurally related vitamins was achieved using a new generation of pentafluorophenyl propyl core-shell stationary phase with elution using methanol and an aqueous solution of ammonium acetate. The fluorescence of retinol and tocopherol isomers was detected at λex = 325, 295 nm and λem = 480, 325 nm, respectively. The rapid baseline separation of all analytes was accomplished within 4.0 min. The sensitivity of method was demonstrated with lower limits of quantification: retinol 0.01 μM, α-tocopherol 0.38 μM, β-tocopherol 0.18 μM, γ-tocopherol 0.14 μM, and δ-tocopherol 0.01 μM. Possible application of this method in clinical practice was confirmed by the analysis of human serum samples from healthy volunteers. Finally, the simultaneous determination of retinol and all tocopherol isomers in human serum can enable the clarification of their role in metabolism and in diseases such as cancer.
A new, rapid and effective ultra-high-performance liquid chromatography method with mass spectrometry detection is described for the separation and quantification of 8-hydroxy-2-deoxyguanosine, 8-hydroxyguanosine and creatinine in human urine. The present study uses an isotope-labelled internal standard ([15N]5-8-hydroxy-2-deoxyguanosine), a BIO core-shell stationary phase and an isocratic elution of methanol and water. Sample preparation of human urine was performed by solid-phase extraction (SPE) on Oasis HLB cartridges with methanol/water 50:50 (v/v) elution. Extraction recoveries ranged from 98.1% to 109.2%. Biological extracts showed high short-term stability. Several aspects of this procedure make it suitable for both clinical and research purposes: a short elution time of less than 3.2 min, an intra-day precision of 2.5-8.9%, an inter-day precision of 3.4-8.7% and low limits of quantification (27.7 nM for 8-hydroxyguanosine, 6.0 nM for 8-hydroxy-2-deoxyguanosine). Finally, simultaneous analysis of DNA and RNA oxidative stress biomarkers is a useful tool for monitoring disease progression in neurodegenerative disorders and cancer. Graphical abstract UHPLC-MS/MS analysis of DNA and RNA oxidative stress biomarkers.
- MeSH
- biologické markery moč MeSH
- deoxyguanosin analogy a deriváty moč MeSH
- DNA moč MeSH
- dospělí MeSH
- extrakce na pevné fázi metody MeSH
- guanosin analogy a deriváty moč MeSH
- kreatin moč MeSH
- lidé MeSH
- limita detekce MeSH
- mladý dospělý MeSH
- nádory moč MeSH
- neurodegenerativní nemoci moč MeSH
- oxidační stres MeSH
- RNA moč MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- validační studie MeSH
Závěrečná zpráva o řešení grantu Interní grantové agentury MZ ČR
1 svazek : ilustrace, tabulky ; 30 cm
Projekt je zaměřen na vývoj a validaci UHPLC metody s využitím nejnovějšího systému UFLC-Nexera pro stanovení vitaminu D (D2 a D3) a jeho metabolitů (25-OH D2, 25-OH D3) v lidském séru s využitím moderních technologií jak v kapalinové chromatografii, tak v úpravě biologického materiálu a přípravě vzorku k analýze. Vývoj metody je iniciován požadavky klinického výzkumu Fakultní nemocnice Hradec Králové (FNHK) . Důraz je kladen nejen na vlastní výzkum studie metabolismu vitaminu D, ale i na rychlost a jednoduchost dané metody, což by umožnilo její zavedení do rutinního vyšetření např. v biochemických laboratořích. Výsledkem této studie by proto měl být přehled o hladinách vitaminu D a jeho metabolitů u vybrané skupiny pacientů. Po vyhodnocení projektu by propojení výzkumných aktivit z oblasti analytické chemie a klinického monitorování mělo vést ke zlepšení diagnostických možností a zároveň zkvalitnění léčebných postupů u závažných civilizačních chorob a zlepšení péče o jednotlivé pacienty.; The project focuses on the development and validation of UHPLC methods for determination of vitamin D and its metabolites (25-OH D2, 25-OH D3) in human serum using modern technology. Method development is initiated by clinical research requirements of the Teaching Hospital in Hradec Kralove. Emphasis is placed not only on own research studies of vitamin D metabolism, but also on the speed and simplicity of the method, which would allow its introduction into routine examinations such as biochemical laboratories. The result of this study should be therefore an overview of the levels of vitamin D and its metabolites in selected groups of patients. After evaluation of the project should link of research activities in the field of analytical chemistry and clinical monitoring lead to better diagnostic capabilities and also improve treatments for serious civilization diseases and improve the care of individual patients.
- MeSH
- biochemická analýza krve MeSH
- diagnostické techniky a postupy MeSH
- hmotnostní spektrometrie MeSH
- hypercholesterolemie diagnóza MeSH
- klinické laboratorní techniky MeSH
- LDL-cholesterol MeSH
- separace krevních složek MeSH
- sérum MeSH
- vitamin D analýza metabolismus MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Konspekt
- Biochemie. Molekulární biologie. Biofyzika
- NLK Obory
- chemie, klinická chemie
- biochemie
- NLK Publikační typ
- závěrečné zprávy o řešení grantu IGA MZ ČR
AIM: To analyze the prognostic significance of serum and urinary neopterin concentrations in patients with rectal adenocarcinoma treated with (chemo)radiation. PATIENTS AND METHODS: Urinary and serum neopterin and peripheral blood cell count were determined in 49 patients with rectal carcinoma before the start of (chemo)radiation. RESULTS: Neopterin concentrations exhibited a significant inverse correlation with hemoglobin and positive correlation with leukocyte count, platelet count and platelet-to-lymphocyte ratio. Increased serum neopterin concentration was associated with significantly inferior relapse-free survival (RFS) and overall survival. However, a significant association was observed only in 28 patients treated in the neoadjuvant setting. Although increased urinary neopterin was also associated with inferior RFS and overall survival, this was not statistically significant. The neutrophil-to-lymphocyte ratio was also associated with poor prognosis. CONCLUSION: The data presented herein indicate a prognostic significance of serum neopterin concentrations in patients with rectal cancer treated with neoadjuvant chemoradiation.
- MeSH
- chemoradioterapie metody MeSH
- lidé středního věku MeSH
- lidé MeSH
- nádory rekta radioterapie terapie MeSH
- neopterin krev moč MeSH
- prognóza MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Vitamin E comprises eight related compounds: α-, β-, γ-, δ-tocopherols and α-, β-, γ-, δ-tocotrienols. In the past, α-tocopherol has been the isomer that was studied most, and its anti-inflammatory and anti-proliferative effects have been described. Therefore, many prevention trials have investigated the effect of α-tocopherol on human health. Current research studies have also defined the important roles of other tocopherols, such as anti-inflammatory, anti-proliferative and cancer preventative effects. Knowledge of the individual tocopherols could help to understand their roles in various metabolic pathways. This review summarizes the recent trends in sample pretreatment, liquid chromatography and selected applications of the determination of tocopherols in various biological materials. The relationship between tocopherol isomers and serious diseases is also described. Graphical Abstract Article structure.
- MeSH
- lidé MeSH
- tělesné tekutiny chemie MeSH
- tokoferoly analýza MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
AIM: A new HPLC method for the determination of neopterin, kynurenine and tryptophan using a second-generation monolith stationary phase and high-throughput sample preparation procedure based on microplates was developed and fully validated. MATERIALS & METHODS: As the stationary phase a monolithic C18 Chromolith high-resolution column with dimensions of 4.6 × 100 mm connected to a monolithic 4.6 × 10-mm security guard was used. Separation was achieved using 15 mM phosphate buffer (KH2PO4 +K2HPO4·3H2O at pH 3) and acetonitrile in gradient mode. RESULTS: Target analytes were determined in 5.5 minutes in amniotic fluid, effusions and wound exudates with a limit of quantification (LOQ) of 1.25 nM for neopterin, 2.5 µM for tryptophan and 0.25 µM for kynurenine. DISCUSSION: The method was applied to real clinical sample measurements, and it will be used to monitor neopterin, kynurenine and tryptophan levels in biological fluids to assess the patient response to therapy and clinical status.
- MeSH
- exsudáty a transsudáty metabolismus MeSH
- hojení ran * MeSH
- kynurenin analýza MeSH
- lidé MeSH
- maligní pleurální výpotek metabolismus MeSH
- neopterin analýza MeSH
- plodová voda metabolismus MeSH
- spektrofotometrie ultrafialová MeSH
- tryptofan analýza MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
