Lipids from microorganisms, and especially lipids from Archaea, are used as taxonomic markers. Unfortunately, knowledge is very limited due to the uncultivability of most Archaea, which greatly reduces the importance of the diversity of lipids and their ecological role. One possible solution is to use lipidomic analysis. Six radioactive sources were investigated, two of which are surface (Wettinquelle and Radonka) and four deep from the Svornost mine (Agricola, Behounek, C1, and Curie). A total of 15 core lipids and 82 intact polar lipids were identified from the membranes of microorganisms in six radioactive springs. Using shotgun lipidomics, typical Archaea lipids were identified in spring water, namely dialkyl glycerol tetraethers, archaeol, hydroxyarchaeol and dihydroxyarchaeol. Diverse groups of polar heads were formed in archaeal IPLs, whose polar heads are formed mainly by hexose, deoxyhexose, and phosphoglycerol. The analysis was performed using shotgun lipidomics and the structure of all molecular species was confirmed by tandem mass spectrometry. After acid hydrolysis, a mixture of polar compounds was obtained from the polar head. Further analysis by GC-MS confirmed that the carbohydrates were glucose and rhamnose. Analysis by HPLC-MS of diastereoisomers of 2-(polyhydroxyalkyl)-3-(O-tolylthiocarbamoyl)thiazolidine-4(R)-carboxylates revealed that both L-rhamnose and D-glucose are present in spring samples only in varying amounts. The glycoside composition depends on the type of spring, that is, Wettinquelle and Radonka springs are basically shallow groundwater, while the samples from the Svornost mine are deep groundwater and do not contain glycosides with rhamnose. This method enables quick screening for characteristic Archaea lipids, allowing decisions on whether to pursue further analyses, such as metagenomic analysis, to directly confirm the presence of Archaea.

• MeSH
• Archaea * chemie   klasifikace   MeSH
• lipidomika * MeSH
• membránové lipidy * chemie   analýza   MeSH
• plynová chromatografie s hmotnostně spektrometrickou detekcí MeSH
• přírodní prameny mikrobiologie   chemie   MeSH
• tandemová hmotnostní spektrometrie MeSH
• Publikační typ
• časopisecké články MeSH

There is growing evidence that endocrine disruptive chemicals have deleterious effects on sexual and reproductive function. To examine subjective sexual functions in human females and their relationship to postnatal phthalate exposure and perinatal androgenization, a Sexuality Score (SS) was established from a first-stage survey questionnaire of subjective sexual function filled out by female university students (n = 68; average age 25.23 ± 5.17 years; rural 25.51 ± 6.74 vs. urban 25.85 ± 1.43 years). Seventeen phthalate metabolites in urine samples were analyzed by high-performance liquid chromatography (HPLC) and tandem mass spectrometry (MS/MS). Females were also assessed for the 2D:4D digit ratio as an index of perinatal androgenization. The mean age of menarche was 12.82 ± 1.35 years (rural 12.59 ± 1.39 vs. urban 13.18 ± 1.27; p = 0.01). The mean age at first sexual intercourse was 14.88 ± 6.89 years (rural 14.62 ± 7.20 vs. urban 15.24 ± 6.55), and as the age of first sexual intercourse increases, the SS score tends to increase as well, albeit moderately (r = 0.25, p = 0.037). Mono-iso-butyl phthalate, mono(2-ethyl-5-carboxypentyl) phthalate, mono(hydroxy-n-butyl) phthalate, mono(2-ethyl-5-oxohexyl) phthalate (p ≤ 0.05) and mono(2-carboxymethylhexyl) phthalate (p ≤ 0.01) were negatively associated with SS. A compounding butterfly effect of prenatal exposure to androgens was observed with disruptive effects of mono(2-ethyl-5-oxohexyl) phthalate and mono(2-ethyl-5-carboxypentyl) phthalate on sexual function. Exposure to phthalates in adult females may lead to disruption of subjective sexual function, especially concerning sexual desire and sexual satisfaction, and perinatal androgenization could augment these effects.

• MeSH
• androgeny * MeSH
• dospělí MeSH
• endokrinní disruptory * MeSH
• kyseliny ftalové * moč   MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• průzkumy a dotazníky MeSH
• sexuální chování * účinky léků   MeSH
• těhotenství MeSH
• zpožděný efekt prenatální expozice * MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Epilepsy, affecting over 50 million people globally, presents a significant neurological challenge. Effective prevention of epileptic seizures relies on proper administration and monitoring of Anti-Seizure Medication (ASMs). Therapeutic Drug Monitoring (TDM) ensures optimal dosage adjustment, minimizing adverse effects and potential drug interactions. While traditional venous blood collection for TDM may be stressful, emerging alternative sampling methods, particularly Dried Blood Spot (DBS) or oral fluid offer less invasive way of sampling. This study aimed to develop and validate an analytical method for the determination of lamotrigine in such alternative samples. The sample, either DBS or oral fluid, was subjected to extraction, evaporation, and reconstitution in 15 % acetonitrile containing 0.1 % formic acid. A Kinetex C18 Polar column was used for liquid chromatographic separation and MS in ESI+ mode was used for detection and quantitation of lamotrigine using an isotopically labelled internal standard according to EMA guidelines. The calibration range of the developed method enables the determination of lamotrigine in the concentration range of 1-30 μg/mL in DBS and 0.5-20 μg/mL in oral fluid. Oral fluid and DBS samples from patients treated with lamotrigine analysed by the developed method were compared to plasma concentrations measured by the hospital's accredited laboratory. Preliminary results indicate a promising potential for these alternative matrices in clinical TDM applications. By offering a less invasive sampling approach, this method improves the accessibility and safety of pharmacotherapy for epilepsy patients. The results of this study lay the foundation for further clinical applications by implementing alternative matrix TDM, which may significantly advance personalized care in epilepsy management.

• MeSH
• antikonvulziva * analýza   krev   MeSH
• chromatografie kapalinová metody   MeSH
• epilepsie farmakoterapie   MeSH
• kalibrace MeSH
• kapalinová chromatografie-hmotnostní spektrometrie MeSH
• lamotrigin * analýza   krev   MeSH
• lidé MeSH
• limita detekce MeSH
• monitorování léčiv * metody   MeSH
• reprodukovatelnost výsledků MeSH
• sliny * chemie   MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• test suché kapky krve * metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• validační studie MeSH

The semi-synthetic cannabinoid hexahydrocannabinol (HHC) has become a highly discussed topic in forensic toxicology since 2022 due to its legal availability at this time and its psychoactive effects. This study aimed to investigate the pharmacokinetics, effects, and immunological detectability of HHC after oral (25 mg HHC fruit gum) and inhalative (three puffs from HHC vape) consumption with three participants per group. Serum (up to 48 h), urine (up to five days), and saliva (up to 48 h) samples were collected at different relevant time points and analyzed by HPLC-MS/MS for (9R)/(9S)-HHC, 11-hydroxy-HHC, and (9R)/(9S)-HHC carboxylic acid with a fully validated method. Additionally, immunological detectability was investigated with three different commercially available tests. To address the psychoactive effects, the subjective "high" feeling (scale 0-10) was monitored and different psychophysical tests (e.g. modified Romberg test, walk and turn) were conducted. Overall, the pharmacokinetics and effects of HHC were comparable to tetrahydrocannabinol (THC). However, the route of administration as well as inter-individual factors played a crucial role regarding maximum concentrations, pharmacokinetic profiles, and psychoactive effects.

• MeSH
• agonisté kanabinoidních receptorů farmakokinetika   farmakologie   MeSH
• aplikace inhalační * MeSH
• aplikace orální * MeSH
• dospělí MeSH
• emoce účinky léků   MeSH
• farmakokinetika * MeSH
• imunologické testy MeSH
• kanabinoidy * analýza   krev   farmakokinetika   farmakologie   moč   MeSH
• kapalinová chromatografie-hmotnostní spektrometrie MeSH
• lidé MeSH
• psychofyziologie * MeSH
• psychotropní léky * analýza   krev   farmakokinetika   farmakologie   moč   MeSH
• sliny chemie   MeSH
• tetrahydrokanabinol farmakokinetika   farmakologie   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

The precise and unambiguous detection and quantification of internal RNA modifications represents a critical step for understanding their physiological functions. The methods of direct RNA sequencing are quickly developing allowing for the precise location of internal RNA marks. This detection is, however, not quantitative and still presents detection limits. One of the biggest remaining challenges in the field is still the detection and quantification of m6A, m6Am, inosine, and m1A modifications of adenosine. The second intriguing and timely question remaining to be addressed is the extent to which individual marks are coregulated or potentially can affect each other. Here, we present a methodological approach to detect and quantify several key mRNA modifications in human total RNA and in mRNA, which is difficult to purify away from contaminating tRNA. We show that the adenosine demethylase FTO primarily targets m6Am marks in noncoding RNAs in HEK293T cells. Surprisingly, we observe little effect of FTO or ALKBH5 depletion on the m6A mRNA levels. Interestingly, the upregulation of ALKBH5 is accompanied by an increase in inosine level in overall mRNA.

• MeSH
• adenosin * analogy a deriváty   metabolismus   genetika   analýza   MeSH
• alfa-ketoglutarát-dependentní dioxygenasa, AlkB homolog 5 * metabolismus   genetika   MeSH
• chromatografie kapalinová metody   MeSH
• gen pro FTO * metabolismus   genetika   MeSH
• HEK293 buňky MeSH
• inosin * metabolismus   genetika   MeSH
• kapalinová chromatografie-hmotnostní spektrometrie MeSH
• lidé MeSH
• messenger RNA * genetika   metabolismus   MeSH
• posttranskripční úpravy RNA MeSH
• tandemová hmotnostní spektrometrie * metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Východiska: Hledání účinných biomarkerů pro včasnou diagnostiku ovariálního karcinomu (ovarian cancer – OC) patří k naléhavým úkolům moderní onkogynekologie. Metabolické profilování pomocí ultra vysokoúčinné kapalinové chromatografie a hmotnostní spektrometrie (ultraigh performance liquid chromatography and mass spectrometry – UHPLC-MS) poskytuje informace o souhrnu všech nízkomolekulárních metabolitů vzorku biologických tekutin pacienta, které odrážejí procesy probíhající v těle. Cílem studie bylo prozkoumat metabolomický profil krevní plazmy a moči pacientek se serózním ovariálním adenokarcinomem pomocí UHPLC-MS. Materiál a metody: K provedení metabolomické analýzy bylo odebráno 60 vzorků krevní plazmy a 60 vzorků moči pacientek s diagnózou serózního karcinomu vaječníků a 20 vzorků zdravých dobrovolníků. Chromatografická separace byla provedena na chromatografu Vanquish Flex UHPLC System (Thermo Scientific, Německo). Analýza hmotnostní spektrometrií byla provedena na Orbitrap Exploris 480 (Thermo Scientific, Německo) vybaveném elektrosprejovým ionizačním zdrojem. Bioinformatická analýza byla provedena pomocí Compound Discoverer Software (Thermo Fisher Scientific, USA), statistická analýza dat byla provedena v programovacím jazyce Python pomocí knihovny SciPy. Výsledky: Pomocí UHPLC-MS bylo v krevní plazmě identifikováno 1 049 metabolitů různých tříd. U pacientek s OC mělo 8 metabolitů významně nižší koncentraci (p < 0,01) ve srovnání se zdravými dárci, zatímco u 19 látek byly zjištěny vyšší hladiny (p < 0,01). Během metabolomického profilování vzorků moči bylo identifikováno 417 metabolitů: 12 látek mělo významně nižší koncentraci ve srovnání se zjevně zdravými jedinci a u 14 látek byly hladiny vyšší (p < 0,01). U pacientek se serózním adenokarcinomem vaječníků byla zjištěna významná změna v metabolomu krevní plazmy a moči, vyjádřená abnormálními koncentracemi lipidů a jejich derivátů, mastných kyselin a jejich derivátů, acylkarnitinů, fosfolipidů, aminokyselin a jejich derivátů, derivátů dusíkatých bází a steroidů. Mezi nejslibnější markery tohoto onemocnění přitom patří kynurenin, kyselina myristová, lysofosfatidylcholin a L-oktanoylkarnitin. Závěr: Odhalené změny v metabolomu se mohou stát základem pro zlepšení přístupů k diagnostice serózního ovariálního adenokarcinomu.

Background: The search for effective biomarkers for ovarian cancer (OC) early diagnosis is an urgent task of modern oncogynecology. Metabolic profiling by ultra-high performance liquid chromatography and mass spectrometry (UHPLC-MS) provides information on the totality of all low molecular weight metabolites of patient’s biological fluids sample, reflecting the processes occurring in the body. The aim of the study was to research blood plasma and urine metabolomic profile of patients with serous ovarian adenocarcinoma by UHPLC-MS. Material and methods: To perform metabolomic analysis, 60 blood plasma samples and 60 urine samples of patients diagnosed with serous ovarian carcinoma and 20 samples of apparently healthy volunteers were taken. Chromatographic separation was performed on a Vanquish Flex UHPLC System chromatograph (Thermo Scientific, Germany). Mass spectrometric analysis was performed on an Orbitrap Exploris 480 (Thermo Scientific, Germany) equipped with an electrospray ionization source. Bioinformatic analysis was performed using Compound Discoverer Software (Thermo Fisher Scientific, USA), statistical data analysis was performed in the Python programming language using the SciPy library. Results: Using UHPLC-MS, 1,049 metabolites of various classes were identified in blood plasma. In patients with OC, 8 metabolites had a significantly lower concentration (P < 0.01) compared with conditionally healthy donors, while the content of 19 compounds, on the contrary, increased (P < 0.01). During the metabolomic profiling of urine samples, 417 metabolites were identified: 12 compounds had a significantly lower concentration compared to apparently healthy individuals, the content of 14 compounds increased (P < 0.01). In patients with ovary serous adenocarcinoma, a significant change in the metabolome of blood plasma and urine was found, expressed in abnormal concentrations of lipids and their derivatives, fatty acids and their derivatives, acylcarnitines, phospholipids, amino acids and their derivatives, derivatives of nitrogenous bases and steroids. At the same time, kynurenine, myristic acid, lysophosphatidylcholine and L-octanoylcarnitine are the most promising markers of this disease. Conclusion: The revealed changes in the metabolome can become the basis for improving approaches to the diagnosis of serous ovarian adenocarcinoma.

• MeSH
• adenokarcinom diagnóza   MeSH
• biologické markery * analýza   krev   moč   MeSH
• kapalinová chromatografie-hmotnostní spektrometrie metody   MeSH
• lidé MeSH
• metabolom * MeSH
• metabolomika metody   MeSH
• nádory vaječníků * diagnóza   MeSH
• ženy MeSH
• Check Tag
• lidé MeSH

While the use of food additives is common manufacturing practice, the levels used in food have to be compliant with the prescribed legislation. For fast control of present levels of food additives in products, ultra-high performance liquid chromatography coupled to tandem mass spectrometry with a triple quadrupole linear ion trap (QTRAP) mass analyser was applied to develop a method for the simultaneous determination of 41 frequently added food additives and flavourings, including 16 water-soluble colourants, 14 illegal dyes, 7 sweeteners, 2 preservatives, and 2 purine alkaloids. The method was validated using energy drink, chilli powder, condiment, and jelly sweets as food sample matrices. The average recovery values were in the range of 70‒120%, and the relative standard deviations were less than 10% for the majority of the analytes. The validated method was applied for the analysis of 134 samples from the Czech market.

• MeSH
• analýza potravin * metody   MeSH
• chuťové esence analýza   MeSH
• kapalinová chromatografie-hmotnostní spektrometrie MeSH
• kontaminace potravin * analýza   MeSH
• lidé MeSH
• limita detekce MeSH
• nápoje * analýza   MeSH
• potravinářské přísady * analýza   MeSH
• reprodukovatelnost výsledků MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• vysokoúčinná kapalinová chromatografie MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• validační studie MeSH
• Geografické názvy
• Česká republika MeSH

Galactooligosaccharides (GOS) are lactose-derived functional ingredients applied in food products and have great potential in health protection. The conversion of lactose to GOS commonly occurs using β-galactosidases of mould, yeast and bacterial origin. The yield and structure of the resulting GOS depend on the enzyme used and the reaction conditions. This work focuses on the structural analysis of the products obtained with four commercial β-galactosidases Maxilact LGI 5000 (ML), Maxilact A4 MG (MA), Saphera 2600 L (SA) and NOLA Fit 5500 (NL) to evaluate their efficiency and specificity. HPLC, ESI-MS and NMR spectroscopy were applied to characterise the GOS preparations. GOS were separated from the reaction mixture using activated charcoal treatment. HPLC analysis confirmed that most of the monosaccharides and a part of the lactose, but also some other disaccharides, probably allolactose and 6-galactobiose, were retained by charcoal. In all the products, ESI-MS analysis detects oligosaccharides up to hexamers. NMR spectra confirmed the presence of GOS of various configurations and polymerisation degrees and evaluated the specificity of used enzymes. MA preferably forms 1,6- and 1,4-glycosidic bonds, and bacterial enzymes NL and SA also form 1,2- and 1,3- glycosidic bonds, while yeast enzyme ML cannot produce new 1,4-glycosidic bonds. The mould enzyme MA showed the highest trans-galactosylation activity, forming longer GOS oligomers than the other enzymes.

• MeSH
• beta-galaktosidasa * metabolismus   chemie   MeSH
• galaktosa chemie   metabolismus   MeSH
• hmotnostní spektrometrie s elektrosprejovou ionizací * metody   MeSH
• laktosa metabolismus   chemie   MeSH
• magnetická rezonanční spektroskopie * metody   MeSH
• oligosacharidy * chemie   metabolismus   MeSH
• vysokoúčinná kapalinová chromatografie metody   MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH

Microflow liquid chromatography interfaced with mass spectrometry (μLC-MS/MS) is increasingly applied for high-throughput profiling of biological samples and has been proven to have an acceptable trade-off between sensitivity and reproducibility. However, lipidomics applications are scarce. We optimized a μLC-MS/MS system utilizing a 1 mm inner diameter × 100 mm column coupled to a triple quadrupole mass spectrometer to establish a sensitive, high-throughput, and robust single-shot lipidomics workflow. Compared to conventional lipidomics methods, we achieve a ∼4-fold increase in response, facilitating quantification of 351 lipid species from a single iPSC-derived cerebral organoid during a 15 min LC-MS analysis. Consecutively, we injected 303 samples over ∼75 h to prove the robustness and reproducibility of the microflow separation. As a proof of concept, μLC-MS/MS analysis of Alzheimer's disease patient-derived iPSC cerebral organoid reveals differential lipid metabolism depending on APOE phenotype (E3/3 vs E4/4). Microflow separation proves to be an environmentally friendly and cost-effective method as it reduces the consumption of harmful solvents. Also, the data demonstrate robust, in-depth, high-throughput performance to enable routine clinical or biomedical applications.

• MeSH
• apolipoproteiny E MeSH
• chromatografie kapalinová metody   MeSH
• fenotyp MeSH
• kapalinová chromatografie-hmotnostní spektrometrie * MeSH
• lidé MeSH
• lipidomika MeSH
• reprodukovatelnost výsledků MeSH
• tandemová hmotnostní spektrometrie * metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The increasing contamination of cereals by micromycetes and mycotoxins during malting still poses an unresolved food safety problem. This study characterises the potential of the novel, rapidly developing food production technology of Pulsed Electric Field (PEF) to reduce the viability of Fusarium fungi and the production of mycotoxins during malting. Barley, artificially inoculated with four Fusarium species, was treated by PEF with two different intensities and then malted using a standard Pilsner-type technology. Concentrations of fungi were quantified by RT-PCR, expression of fungal growth-related genes was assessed using mRNA sequencing, and mycotoxin levels were analysed by U-HPLC-HRMS/MS. Despite the different trends for micromycetes and mycotoxins after application of variously intense PEF conditions, significant reductions were generally observed. The greatest decrease was for F. sporotrichioides and F. poae, where up to six fold lower levels were achieved for malts produced from the PEF-treated barley when compared to the control. For F. culmorum and F. graminearum, up to a two-fold reduction in the PEF-generated malts was observed. These reductions mostly correlated with a decrease in relevant mycotoxins, specifically type A trichothecenes.

• MeSH
• elektřina MeSH
• Fusarium * metabolismus   genetika   růst a vývoj   MeSH
• ječmen (rod) * mikrobiologie   MeSH
• jedlá semena * mikrobiologie   MeSH
• kontaminace potravin prevence a kontrola   MeSH
• manipulace s potravinami metody   MeSH
• mykotoxiny * biosyntéza   metabolismus   MeSH
• potravinářská mikrobiologie MeSH
• Publikační typ
• časopisecké články MeSH