Tomaž Vovk

Supportive therapy with haemoadsorption is gaining popularity in critically ill patients, with the aim of reducing overinflammation triggered by the cytokine storm. The haemoadsorbers used are not specific for cytokines and also bind antibiotics. The aim of this study was to develop and validate a simple analytical method for the simultaneous determination of selected antibiotics and to develop an in vitro model for the quantification of their binding to the CytoSorb^® haemoadsorber under conditions simulating sepsis. Imipenem (IMI), amoxicillin (AMO), cefepime (CEF), meropenem (MERO), vancomycin (VAN) and piperacillin (PIP) were measured in bovine plasma via precipitation with acetonitrile and liquid–liquid extraction with dichloromethane. The aqueous phase was collected and analysed using a C18 HPLC system under gradient conditions, with modulation of organic solvent content and mobile phase pH, and detection performed using a UV/Vis detector. The method was linear (r² > 0.982) across investigated analytical ranges (1.0–100.0 µg/mL for AMO and VAN, 1.0–75.0 µg/mL for CEF, MERO and PIP and 2.5–100.0 µg/mL for IMI). Intra- and inter-day precision did not exceed 14% and accuracy ranged from 85.8% to 108.5%. Using the in vitro model, we showed that CytoSorb^® significantly removed VAN and PIP, but not MERO. Further clinical studies are needed to establish the clinical significance of these findings and their impact on antibiotic exposure.

Background/Objectives: Haemoadsorption has recently emerged as an extracorporeal treatment option for sepsis, septic shock, intoxications, and cardiac surgery to modulate dysregulated inflammatory responses or remove a wide range of circulating molecules. To ensure appropriate clinical use of the CytoSorb^® haemoadsorber, it is essential to understand the extent to which specific drugs are adsorbed by the device. Methods: We conducted a systematic literature review using the PubMed and Ovid MEDLINE database to identify studies on drug binding to the CytoSorb^® haemoadsorber, including both in vivo and in vitro studies. Publications in English language, available up to 31 December 2025 that reported or enabled calculation of percentage of drug removal, CytoSorb^® clearance or half-life during CytoSorb^® therapy were included. Records were screened, eligibility and quality were assessed, and data were extracted independently by two reviewers. Results: We found that 26 studies reported on the binding of 56 drugs to CytoSorb^®, with most available information relating to antibiotics used in the treatment of sepsis and septic shock. CytoSorb^® appears to remove vancomycin and linezolid but not meropenem, although data for other antibiotics are insufficient to assess clinical relevance. Data on the removal of anticoagulant and antithrombotic drugs with CytoSorb^® before and during cardiac surgery indicate that using this procedure to reduce complications associated with apixaban and ticagrelor is feasible and safe. The available evidence on the use of CytoSorb^® for drug poisoning is of very low quality. Conclusions: Although the number of studies on drug binding to the CytoSorb^® is increasing, the review is limited by the marked heterogeneity among the included studies. It is advised to use therapeutic drug monitoring whenever possible during CytoSorb^® treatment. Research of binding of drugs to CytoSorb^® is crucial for its safe and effective clinical use, but adequate methodology is necessary.

Background/Objectives: Dried blood spot (DBS) sampling, a technique for collecting capillary blood samples, is widely used in therapeutic drug monitoring, pharmacokinetic and toxicology research, newborn screening, and population health because it enables simple, non-invasive sampling across large cohorts. However, it presents several challenges, mainly due to the effect of hematocrit (HCT), which can influence the quantification of analytes. Methods: A combination of methods was developed to estimate the HCT and blood volume in DBS samples. Image analysis and hemoglobin (Hb) quantification using UV-VIS spectrometry were used for HCT estimation, and conductivity was used to determine blood volume. DBS samples from five donors were prepared with HCT between 0.2 and 0.6 and were used to prepare calibrators and quality control samples. The developed methods were applied to 23 samples obtained from ten adult patients with inflammatory bowel disease. Results: The methods for HCT determination using image analysis or Hb measurements were linear (r² > 0.994), with acceptable accuracy (90.3–102.2%) and precision (2 = 0.999) and enabled accurate (96.8–100%) and precise (<5.65%) determination of blood volume in DBS samples. All three methods were in good agreement with the reference values in patient samples. Finally, strategies to correct HCT- and volume-related bias in DBS samples were proposed for analytes with different blood cell-to-plasma partition coefficients. Conclusions: We accurately and precisely estimated HCT in DBS samples using image analysis and Hb determination, and the volume of blood in DBS using conductivity measurement. We evaluated different approaches and derived an optimal procedure for HCT-bias correction.

Lumateperone is a recently approved atypical antipsychotic for the treatment of schizophrenia, with moderate adverse effects. However, the risk of toxicity, mainly due to possible multidrug coadministration, makes the therapeutic drug monitoring (TDM) approach crucial for developing personalised therapies and reducing side effects. In the TDM research field, microsampling represents an innovative and effective approach based on the collection and analysis of small sample volumes (< 100 μL), exhibiting many advantages (such as enhanced analyte stability, minimally invasive procedures and simplified sample storage and transport) and improving patient adherence. For the first time, an analytical methodology based on microsampling was developed and fully validated for the analysis of lumateperone in capillary blood microsamples, by means of volumetric absorptive microsampling (VAMS) in comparison with plasma analysis, with the future perspective of supporting the TDM of psychiatric patients. A reliable and streamlined VAMS protocol was optimised, and an original HPLC–MS/MS method was developed and validated, granting satisfactory results on simulated samples in terms of linearity, precision, extraction yield and matrix effect. Method reliability was successfully tested by comparing the VAMS performance with standard in-tube fluid plasma, and short- and medium-term comparative stability assays confirmed the enhancement of analyte stability obtained in dried microsamples with respect to plasma samples. This is the first analytical method based on microsampling able to provide reliable data when compared to plasma analysis and promising for future TDM applications in patients treated with lumateperone, thus offering significant advantages over conventional blood/plasma sampling in terms of collection, storage and processing. Graphical abstract

Oxidative stress is increasingly recognized as a significant contributor to the pathogenesis of inflammatory bowel disease (IBD). However, the clinical utility of oxidative stress-related biomarkers for assessing and predicting disease activity remains unclear. This prospective observational study aimed to evaluate the diagnostic and predictive performance of oxidative stress-related biomarkers in distinguishing between active IBD and remission across clinical, biochemical, and endoscopic criteria. A total of 76 patients with IBD were followed across three visits: baseline (biological treatment initiation), post-induction (6–12 weeks), and final follow-up (24–36 weeks). Associations with clinical, biochemical, and endoscopic remission status at the final follow-up were evaluated using correlation matrices, receiver operating characteristic curve analysis, principal component analysis, and logistic regression. Ceruloplasmin, plasma free thiols, gamma-glutamyl transferase, and albumin showed significant diagnostic values for distinguishing active disease from remission, using C-reactive protein (CRP)-based criteria. Serum uric acid, advanced oxidation protein products, gamma-glutamyl transferase, total antioxidant capacity, and ceruloplasmin predicted clinical or CRP-based remission when measured at baseline or post-induction, with predictive value varying by biomarker and the time point. Overall, our findings reinforce the role of oxidative stress in the pathogenesis of IBD and highlight the potential of oxidative stress-related biomarkers to be used as tools for monitoring disease activity and predicting IBD treatment outcomes.

Background: Ustekinumab (UST) is a monoclonal antibody (mAb) used in the treatment of inflammatory bowel disease (IBD). Elevated serum concentrations are typically associated with improved therapeutic outcomes, and therapeutic drug monitoring (TDM) is a useful tool for guiding mAbs treatment. This study aimed to develop a dried blood spot (DBS) method for TDM of UST in patients with IBD. Methods: The commercial enzyme-linked immunosorbent assay for plasma samples was optimized for DBS samples and subsequently validated according to international guidelines for classical and DBS-specific validation parameters. It was then applied to analyze serum and DBS samples obtained from venous and capillary blood of IBD patients undergoing UST therapy. Results: The method was linear (3–12 mg/L) with acceptable inter-day accuracy (90.1–106%) and precision (<12%). We confirmed that there was no hematocrit effect and that DBS samples were stable for one month under room conditions. A linear model was developed between venous DBS and serum UST concentrations, which showed no systemic bias, and 71% of the samples were within ±20% of the mean. In addition, a linear correlation between venous DBS and capillary DBS samples was established, showing no significant bias, with 84% of samples within ±20% of the mean. Finally, a novel strategy was developed to overcome the limitations of poor-quality samples (irregular shapes) based on area image analysis. Conclusions: The newly developed DBS method is the first to enable reliable measurement of UST in capillary blood, appropriate clinical interpretation of the measured concentrations, and remote monitoring of patients in the early phase of therapy.

Background/Objectives: Vancomycin, a glycopeptide antibiotic used for gram-positive infections, is associated with acute kidney injury (AKI). Therapeutic drug monitoring (TDM) is recommended to minimize this risk while ensuring therapeutic efficacy. This study evaluated whether AUC-guided monitoring improved patient safety compared to traditional trough-guided monitoring. Methods: A retrospective observational cohort study was conducted at the University Medical Centre Maribor, Slovenia, involving patients receiving intravenous vancomycin. One cohort was managed using trough-guided monitoring (n = 85), while the other was monitored using the AUC-guided approach (n = 139). The primary outcome was AKI incidence, and secondary outcomes included renal replacement therapy and mortality. Risk factors for AKI were identified, and pharmacokinetic parameters were evaluated at vancomycin therapy initiation and steady state. Results: The incidence of AKI was 20% in the trough-guided group and 18% in the AUC-guided group (p = 0.727). Secondary outcomes were similar in both cohorts. Risk factors for AKI included older age (OR 1.04; p = 0.042), higher steady-state AUC (OR 1.01; p < 0.001), longer duration of concomitant nephrotoxic therapy (OR 1.06; p = 0.019), and concomitant use of loop diuretics (OR 2.46; p = 0.045). Steady-state AUC values and trough levels (AUC_0–24ss, AUC_24–48ss, AUC_0–48ss, and C_min48ss) were significantly lower in the AUC-guided group, which was further reflected in the lower percentage of patients exceeding the AUC > 600 mg·h/L threshold at steady state. Conclusions: Although AKI incidence was lower in the AUC-guided group, the difference did not reach statistical significance. However, lower AUC values and trough levels in the AUC-guided group at steady state suggest a trend toward reduced vancomycin exposure and toxicity.

Extracellular nanoparticles (EPs) are a subject of increasing interest for their biological role as mediators in cell–cell communication; however, their harvesting and assessment from bodily fluids are challenging, as processing can significantly affect samples. With the aim of minimizing processing artifacts, we assessed the number density (n) and hydrodynamic diameter (D_h) of EPs directly in diluted plasma and blood using the following recently developed technique: interferometric light microscopy (ILM). We analyzed 613 blood and plasma samples from human patients with inflammatory bowel disease (IBD), collected in trisodium citrate and ethylenediaminetetraacetic acid (EDTA) anticoagulants, and 163 blood and plasma samples from canine patients with brachycephalic obstructive airway syndrome (BOAS). We found a highly statistically significant correlation between n in the plasma and n in the blood only in the human (i.e., but not canine) blood samples, between the samples with trisodium citrate and EDTA, and between the respective D_h for both species (all p < 10⁻³). In the human plasma, the average <D_h> was 139 ± 31 nm; in the human blood, <D_h> was 158 ± 11 nm; in the canine plasma, <D_h> was 155 ± 32 nm; and in the canine blood, <D_h> was 171 ± 33 nm. The differences within species were statistically significant (p < 10⁻²), with sufficient statistical power (P > 0.8). For <n>, we found no statistically significant differences between the human plasma and blood samples or between the samples with trisodium citrate and EDTA. Our results prove that measuring n and D_h of EPs in minimally processed fresh blood and in diluted fresh plasma by means of ILM is feasible for large populations of samples.

Background and Objectives: Oxidative stress resulting from a disturbance of the endogenous redox system is suspected in numerous diseases of the central nervous system, including epilepsy. In addition, antiseizure medications (ASMs), especially those of the old generation, may further increase oxidative stress. To evaluate the effects of ASM generation on oxidative stress, we conducted a cross-sectional study in patients with epilepsy treated with old, new, and polytherapy. Materials and Methods: The antioxidant activity of superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, as well as the concentrations of malondialdehyde, protein carbonyl, nitrate, nitrite, and glutathione in reduced and oxidized forms, were measured in 49 patients with epilepsy and 14 healthy controls. In addition, the plasma concentrations of ASMs and metabolites of carbamazepine and valproic acid were measured in the patients. Results: Patients with epilepsy showed increased activities of superoxide dismutase and catalase (p < 0.001), concentrations of glutathione disulfide and markers of nitric oxide metabolism (p < 0.001), and decreased activities of glutathione peroxidase, glutathione reductase, glutathione, and nitrite concentrations (p ≤ 0.005) compared to healthy controls. A comparison of ASM generations revealed increased levels of superoxide dismutase and catalase (p ≤ 0.007) and decreased levels of glutathione peroxidase and glutathione reductase (p ≤ 0.01) in patients treated with old ASMs compared to those treated with new generation ASMs. In addition, an increase in protein carbonyl and nitric oxide metabolites (p ≤ 0.002) was observed in patients treated with old generation ASMs compared to those treated with new generation ASMs. Most oxidative stress parameters in patients receiving polytherapy with ASMs were intermediate between the results of patients treated with the old and new generations of ASMs. Conclusions: An increase in oxidative stress markers and modulation of antioxidant enzyme activities was observed in patients with epilepsy compared to controls. The results of our study showed significantly higher oxidative stress in patients treated with old ASMs compared to those treated with new generation ASMs.