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Due to the emergence of novel psychoactive substances on the drug market, there is a growing demand for analytical methods allowing identification and determination of as many psychoactive substances as possible in the shortest possible time, which can be easily expanded to include additional analytes appearing in street trade. Immunochemical methods are not sufficient to meet constantly growing requirements. Therefore, the aim of the study was to develop an analytical method enabling quick analysis of urine samples for psychoactive substances, drugs and their metabolites. Liquid-liquid extraction (LLE) and liquid chromatography coupled with mass spectrometry (LC-MS/MS) were used for this purpose. Using available analytical standards, the operating parameters of the mass spectrometer were selected for each of the 477 analytes, and MRM (multiple reaction monitoring) acquisition was selected for each of them. The use of analytical standards allowed for the identification of analytes separated on the chromatography column. The exceptions are methylmethcathinone isomers (3-MMC and 4-MMC), which we were unable to separate using the gradient elution method used. Extraction using acetonitrile with the addition of ammonium formate and formic acid allowed us to obtain high recoveries without the use of β-glucuronidase. Recovery values ranged from 18.43 to 119.94%. The matrix effect was eliminated by obtaining a calibration curve in the matrix. The developed analytical method was validated in accordance with SWGTOX guidelines. Only 12 substances did not meet the validation criteria (CV: ±20% and bias: ±20%). Thus the validated method makes it possible to determine 465 psychoactive substances in just 30 minutes. In the validation process, values such as the limit of detection (LOD) and the limit of quantification (LOQ) were also determined. The LODs are in the range of 3-30 ng mL -1 , and the LOQs are in the range of 10-100 ng mL -1 . The method was successfully applied to toxicological analyses of urine samples, which was an opportunity to develop it further to meet the needs of toxicology.

Endometrial cancer (EC) accounts for 90% of uterine cancer cases. It is considered not only one of the most common gynecological malignancies but also one of the most frequent cancers among women overall. Nowadays, the differentiation of EC subtypes is based on immunohistochemistry and molecular techniques. It is considered that patients' prognosis and the implementation of the appropriate treatment depend on the cancer subtype. Patients with pathogenic variants in POLE have the most favorable outcome, while those with abnormal p53 protein have the poorest. Therefore, in patients with POLE mutation, the de-escalation of postoperative treatment may be considered, and patients with abnormal p53 protein should be subjected to intensive adjuvant therapy. Patients with a DNA mismatch repair (dMMR) deficiency are classified in the intermediate prognosis group as EC patients without a specific molecular profile. Immunotherapy has been recognized as an effective treatment method in patients with advanced or recurrent EC with a mismatch deficiency. Thus, different adjuvant therapy approaches, including targeted therapy and immunotherapy, are being proposed depending on the EC subtype, and international guidelines, such as those published by ESMO and ESGO/ESTRO/ESP, include recommendations for performing the molecular classification of all EC cases. The decision about adjuvant therapy selection has to be based not only on clinical data and histological type and stage of cancer, but, following international recommendations, has to include EC molecular subtyping. This review describes how molecular classification could support more optimal therapeutic management in endometrial cancer patients.

Despite the different possible paths of treatment, lung cancer remains one of the leading causes of death in oncological patients. New tools guiding the therapeutic process are under scientific investigation, and one of the promising indicators of the effectiveness of therapy in patients with NSCLC is variant allele frequency (VAF) analysis. VAF is a metric characterized as the measurement of the specific variant allele proportion within a genomic locus, and it can be determined using methods based on NGS or PCR. It can be assessed using not only tissue samples but also ctDNA (circulating tumor DNA) isolated from liquid biopsy. The non-invasive characteristic of liquid biopsy enables a more frequent collection of material and increases the potential of VAF analysis in monitoring therapy. Several studies have been performed on patients with NSCLC to evaluate the possibility of VAF usage. The research carried out so far demonstrates that the evaluation of VAF dynamics may be useful in monitoring tumor progression, remission, and recurrence during or after treatment. Moreover, the use of VAF analysis appears to be beneficial in making treatment decisions. However, several issues require better understanding and standardization before VAF testing can be implemented in clinical practice. In this review, we discuss the difficulties in the application of ctDNA VAF analysis in clinical routine, discussing the diagnostic and methodological challenges in VAF measurement in liquid biopsy. We highlight the possible applications of VAF-based measurements that are under consideration in clinical trials in the monitoring of personalized treatments for patients with NSCLC.

Dietary supplements are widely available products used by millions of people around the world. Unfortunately, the procedure of adding pharmaceutical and psychoactive substances has recently been observed, in order to increase the effectiveness of supplements in the form of hemp oils. For this reason, it is extremely important to develop analytical methods for the detection of substances prohibited in dietary supplements and food products. In the present study, using the LC-MS/MS technique, an innovative method for the detection and quantification of 117 synthetic cannabinoids and 13 natural cannabinoids in dietary supplements and food products in the form of oils during one 13-min chromatographic run was developed. Each method was fully validated by characterization of the following parameters: The limit of detection was set to 0.1 ng/mL (100 µg/g, 0.01%). The limit of quantification ranged from 0.05 ng/mL to 50 ng/mL. The criteria assumed for systematic error caused by methodological bias (±20%) resulting from the recovery of analytes after the extraction process, as well as the coefficient of variation (CV) (≤20%), were met for all 130 tested compounds. The positive results of the validation confirmed that the developed methods met the requirements related to the adequacy of their application in a given scope. Additionally, methods developed using the LC-MS/MS technique were verified via proficiency tests. The developed analytical procedure was successfully used in the analysis of hemp oils and capsules containing them in the studied dietary supplements.

The emergence of new psychoactive substances on the market is a significant problem on a global scale. This type of substance in society is associated with many negative consequences, such as traffic accidents, accidents at work, rape, homicide, poisoning, or overdose deaths. The analysis of these substances in biological samples is very important for further legal action and saving lives. Therefore, laboratories face a tremendous challenge in tackling the evolving drug market. The paper describes the optimization of the analytical LC-MS/MS method to identify and determine 513 psychoactive substances in hair samples. A method of chromatographic separation was developed, and the working parameters of the mass spectrometer were selected for each analyte. The method has been validated, and the results are as follows: the limit of quantification of the developed method ranges from 0.025 to 1.25 ng/mg hair. The mean recovery of the tested analytes ranges from 80 to 120%. The achieved coefficient of variation in within-run precision ranged from 1.05 to 19.99%. The results achieved for BIAS are in the range of ± 20%., (© 2022. The Author(s).)

Background: Impairments in various subdomains of memory have been associated with chronic cannabis use, but less is known about their neural underpinnings, especially in the domain of the brain's oscillatory activity. Aims: To investigate neural oscillatory activity supporting working memory (WM) in regular cannabis users and non-using controls. We focused our analyses on frontal midline theta and posterior alpha asymmetry as oscillatory fingerprints for the WM's maintenance process. Methods: 30 non-using controls (CG) and 57 regular cannabis users-27 exclusive cannabis users (CU) and 30 polydrug cannabis users (PU) completed a Sternberg modified WM task with a concurrent electroencephalography recording. Theta, alpha and beta frequency bands were examined during WM maintenance. Results: When compared to non-using controls, the PU group displayed increased frontal midline theta (FMT) power during WM maintenance, which was positively correlated with RT. The posterior alpha asymmetry during the maintenance phase, on the other hand, was negatively correlated with RT in the CU group. WM performance did not differ between groups. Conclusions: Both groups of cannabis users (CU and PU), when compared to the control group, displayed differences in oscillatory activity during WM maintenance, unique for each group (in CU posterior alpha and in PU FMT correlated with performance). We interpret those differences as a reflection of compensatory strategies, as there were no differences between groups in task performance. Understanding the psychophysiological processes in regular cannabis users may provide insight on how chronic use may affect neural networks underlying cognitive processes, however, a polydrug use context (i.e., combining cannabis with other illegal substances) seems to be an important factor., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Binkowska, Jakubowska, Krystecka, Galant, Piotrowska-Cyplik and Brzezicka.)

Background While research has consistently identified an association between long-term cannabis use and memory impairments, few studies have examined this relationship in a polydrug context (i.e., when combining cannabis with other substances). Aims: In this preliminary study, we used event-related potentials to examine the recognition process in a visual episodic memory task in cannabis users (CU) and cannabis polydrug users (PU). We hypothesized that CU and PU will have both-behavioral and psychophysiological-indicators of memory processes affected, compared to matched non-using controls with the PU expressing more severe changes. Methods 29 non-using controls (CG), 24 CU and 27 PU were enrolled into the study. All participants completed a visual learning recognition task while brain electrical activity was recorded. Event-related potentials were calculated for familiar (old) and new images from a signal recorded during a subsequent recognition test. We used receiver operating characteristic curves for behavioral data analysis. Results The groups did not differ in memory performance based on receiver operating characteristic method in accuracy and discriminability indicators nor mean reaction times for old/new images. The frontal old/new effect expected from prior research was observed for all participants, while a parietal old/new effect was not observed. While, the significant differences in the late parietal component (LPC) amplitude was observed between CG and PU but not between CG and CU nor CU and PU. Linear regression analysis was used to examine the mean amplitude of the LPC component as a predictor of memory performance accuracy indicator. LPC amplitude predicts recognition accuracy only in the CG. Conclusion The results showed alterations in recognition memory processing in CU and PU groups compared to CG, which were not manifested on the behavioral level, and were the most prominent in cannabis polydrug users. We interpret it as a manifestation of the cumulative effect of multiple drug usage in the PU group., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Binkowska, Jakubowska, Gaca, Galant, Piotrowska-Cyplik and Brzezicka.)

The aim of the study was to evaluate the effect of herbicidal ionic liquids on the population changes of microorganisms used in a batch anaerobic digester. The influence of the following ionic liquids: benzalkonium (2,4-dichlorophenoxy)acetate (BA)(2,4-D), benzalkonium (4-chloro-2-methylphenoxy)acetate (BA)(MCPA), didecyldimethylammonium (2,4-dichlorophenoxy)acetate (DDA)(2,4-D), didecyldimethylammonium (4-chloro-2-methylphenoxy)acetate (DDA)(MCPA), as well as reference herbicides (4-chloro-2-methylphenoxy)acetic acid (MCPA) and (2,4-dichlorophenoxy)acetic acid (2,4-D) in the form of sodium salts on biogas production efficiency was investigated. The effective concentration (EC50) values were determined for all tested compounds. (MCPA) - was the most toxic, with an EC50 value of 38.6⁻41.2 mg/L. The EC50 for 2,4-D was 55.7⁻59.8 mg/L. The addition of the test substances resulted in changes of the population structure of the microbiota which formed the fermentation pulp. The research was based on 16S rDNA analysis with the use of the Next Generation Sequencing method and the MiSeq platform (Illumina, San Diego, CA, USA). There was a significant decrease in bacteria belonging to Firmicutes and Archaea belonging to Euryarchaeota . A significant decrease of the biodiversity of the methane fermentation microbiota was also established, which was expressed by the decrease of the operational taxonomic units (OTUs) and the value of Shannon's entropy. In order to determine the functional potential of bacterial metapopulations based on the 16SrDNAprofile, the PICRUSt(Phylogenetic Investigation of Communities by Reconstruction of Unobserved States)tool was used, which allowed to determine the gene potency of microorganisms and their ability to biodegrade the herbicides. In the framework of the conducted analysis, no key genes related to the biodegradation of MCPA or 2,4-D were found, and the observed decrease of their content in the supernatant liquid was caused by their sorption on bacterial biomass.