In Vitro and In Silico of Cholinesterases Inhibition and In Vitro and In Vivo Anti-Melanoma Activity Investigations of Extracts Obtained from Selected Berberis Species.

Berberis species have a long history of use in traditional Chinese medicine, Ayurvedic medicine, and Western herbal medicine. The aim of this study was the quantification of the main isoquinoline alkaloids in extracts obtained from various Berberis species by HPLC, in vitro and in silico determination of anti-cholinesterase activity, and in vitro and in vivo investigations of the cytotoxic activity of the investigated plant extracts and alkaloid standards. In particular, Berberis species whose activity had not been previously investigated were selected for the study. In the most investigated Berberis extracts, a high content of berberine and palmatine was determined. Alkaloid standards and most of the investigated plant extracts exhibit significant anti-cholinesterase activity. Molecular docking results confirmed that both alkaloids are more favourable for forming complexes with acetylcholinesterase compared to butyrylcholinesterase. The kinetic results obtained by HPLC-DAD indicated that berberine noncompetitively inhibited acetylcholinesterase, while butyrylcholinesterase was inhibited in a mixed mode. In turn, palmatine exhibited a mixed inhibition of acetylcholinesterase. The cytotoxic activity of berberine and palmatine standards and plant extracts were investigated against the human melanoma cell line (A375). The highest cytotoxicity was determined for extract obtained from Berberis pruinosa cortex. The cytotoxic properties of the extract were also determined in the in vivo investigations using the Danio rerio larvae xenograft model. The obtained results confirmed a significant effect of the Berberis pruinosa cortex extract on the number of cancer cells in a living organism. Our results showed that extracts obtained from Berberis species, especially the Berberis pruinosa cortex extract, can be recommended for further in vivo experiments in order to confirm the possibility of their application in the treatment of neurodegenerative diseases and human melanoma.

MOF-801 based solid phase microextraction fiber for the monitoring of indoor BTEX pollution.

Benzene, toluene, ethylbenzene and xylenes (BTEX) are some of the better-known indoor air pollutants, for which effective monitoring is important. The analysis of BTEX can be performed by different type of solid phase microextraction (SPME) fibers. This study presents a proposal for a low cost, convenient and environmentally friendly analytical method for the determination of BTEX in air samples using custom made SPME fibers. In this context, custom made metal organic frameworks (MOF-801) were coated on a stainless-steel wire for SPME fiber preparation. The analysis of BTEX was performed by introducing SPME fiber into an analyte-containing Tedlar bag in steady-state conditions. After the sampling step, the analytes were analyzed using gas chromatography mass spectrometry in selected ion monitoring mode. Parameters that affect the analysis results were optimized; these include desorption temperature and time, preconditioning time, extraction temperature and time, and sample volume. Under optimized conditions, analytical figure of merits of developed method were obtained, including limits of detection (LOD) (0.012 - 0.048 mg/m3), linear ranges (0.041-18 mg/m3), intraday and interday repeatability (2.08 - 4.04% and 3.94 - 6.35%), and fiber to fiber reproducibility (7.51 - 11.17%). The proposed method was successfully applied to real air samples with an acceptable recovery values between 84.5% and 110.9%. The developed method can be applied for the effective monitoring of BTEX.

Comparison of supercritical CO2 extraction and pressurized fluid extraction for isolation of alkaloids from Anacardium occidentale with the study of its anti-inflammatory activity.

In recent years, there has been a growing interest in the therapeutic potential of natural compounds, particularly of plant origin, owing to their demonstrated anti-inflammatory properties. Among these, Anacardium occidentale, commonly known as cashew, has garnered significant attention due to its reputed health benefits. This study aim to establish a correlation between the bioactive compounds contained in the extracts of Anacardium occidentale and its anti-inflammatory activity. Dried Anacardium occidentale leaves powder was used as the extraction matrix. Extraction techniques are maceration, pressurized fluid extraction (PFE), and supercritical fluid extraction (SFE). The preliminary analysis of extracts was made by LC-MS/MS. The Response Surface Methodology (RSM), Principal Component Analysis (PCA), and heat maps were employed to model the influence of experimental conditions on extraction yield and peak area of specific compounds from the plant. To evaluate anti-inflammatory activity, RAW 264.7 cells were cultured, activated with LPS, and treated with varying concentrations of the plant extracts. Cell proliferation was assessed using the XTT assay. Indeed, Anacardium occidentale extracts contain anacardic acids, cardanols, and cardol, with distinct profiles yielded by SFE and ethanol-based methods. RSM shows that temperature and ethanol, as additives to CO2, significantly affect extraction efficiency in both PFE and SFE. Moreover, this composition with SFE demonstrate higher selectivity for specific group of compounds. The extracts exhibit anti-inflammatory properties without cytotoxicity in macrophages, reducing levels of pro-inflammatory proteins COX-2, COX-1, and TLR4 in activated cells. This suggests their potential as anti-inflammatory agents without adverse effects on cell viability or pro-inflammatory protein levels in non-activated cells. Overall, these findings underscore the promising therapeutic potential of Anacardium occidentale extracts in mitigating inflammation, while also providing crucial insights into optimizing the extraction process for targeted compound isolation. Thus, this makes a good prospect for the development of anti-inflammatory drugs from this plant.

Retention mechanism on phosphodiester stationary phases in hydrophilic interaction liquid chromatography and purely aqueous mobile phase part II: Overloading with limited soluble samples.

The adsorption behaviour of caffeine and theophylline under hydrophilic interaction chromatography and purely aqueous conditions was investigated on four phosphodiester stationary phases. Solute adsorption isotherms were determined by frontal analysis or inverse method. The bi-Langmuir model was found to be the best choice to describe the behaviour of caffeine and theophylline adsorption in purely aqueous conditions, whereas the bi-Moreau model describes the adsorption phenomena in HILIC conditions. The results obtained demonstrate that the interaction of caffeine and theophylline with the stationary phase surface varies depending on the mobile phase composition. Both in pure aqueous mobile phase and in HILIC mode, the heterogeneity of the surface of the studied stationary phases is confirmed. In hydrophilic solutions, the sample molecules interact with the stationary phase only. In hydrophobic conditions, a lateral interaction occurs between caffeine or theophylline molecules, which are poorly soluble in acetonitrile-rich solvents. This confirms that the same compound on the same stationary phase can behave rather differently, depending on the mobile phase composition. Thus, the mobile phase may govern and control the retention mechanism.

Development and Validation of LC-MS/MS Method for Determination of Cytisine in Human Serum and Saliva.

Cytisine (CYT) is a quinolizidine alkaloid used for nicotine addiction treatment. Recent clinical trial data regarding cytisine confirm its high effectiveness and safety as a smoking cessation treatment. CYT's popularity is growing due to its increased availability and licensing in more countries worldwide. This increased use by smokers has also resulted in an urgent need for continued drug research, including developing appropriate analytical methods for analyzing the drug in biological samples. In this study, a simple, fast, and reliable method combining hydrophilic interaction liquid chromatography and electrospray ionization quadrupole time of flight mass spectrometry (HILIC/ESI-QTOF-MS) for the determination of CYT in human serum and saliva was developed and validated. This was undertaken after the previous pre-treatment of the sample using solid-phase extraction (SPE). A hydrophilic interaction liquid chromatography (HILIC) column with a silica stationary phase was used for chromatographic analysis. In a linear gradient, the mobile phase consisted of acetonitrile (ACN) and formate buffer at pH 4.0. The proposed method was fully validated and demonstrated its sensitivity, selectivity, precision, and accuracy. The method was successfully applied to determine CYT in serum and, for the first time, in saliva. The findings indicate that saliva could be a promising non-invasive alternative to measure the free concentration of CYT.

Immobilized enzyme microreactors for analysis of tryptic peptides in ß-casein and ß-lactoglobulin.

In this study, our primary objective was to develop an effective analytical method for studying trypsin-digested peptides of two proteins commonly found in cow's milk: ß-casein (ßCN) and ß-lactoglobulin (ßLG). To achieve this, we employed two distinct approaches: traditional in-gel protein digestion and protein digestion using immobilized enzyme microreactors (µ-IMER). Both methods utilized ZipTip pipette tips filled with C18 reverse phase media for sample concentration. The µ-IMER was fabricated through a multi-step process that included preconditioning the capillary, modifying its surface, synthesizing a monolithic support, and further surface modification. Its performance was evaluated under HPLC chromatography conditions using a small-molecule trypsin substrate (BAEE). Hydrolysates from both digestion methods were analyzed using MALDI-TOF MS. Our findings indicate that the µ-IMER method demonstrated superior sequence coverage for oxidized molecules in ßCN (33 ± 1.5%) and ßLG (65 ± 3%) compared to classical in-gel digestion (20 ± 2% for ßCN; 49 ± 2% for ßLG). The use of ZipTips further improved sequence coverage in both classical in-gel digestion (26 ± 1% for ßCN; 60 ± 4% for ßLG) and µ-IMER (41 ± 3% for ßCN; 80 ± 5% for ßLG). Additionally, phosphorylations were identified. For ßCN, no phosphorylation was detected using classical digestion, but the use of ZipTips showed a value of 27 ± 4%. With µ-IMER and µ-IMER-ZipTip, the values increased to 30 ± 2% and 33 ± 1%, respectively. For ßLG, the use of ZipTip enabled the detection of a higher percentage of modified peptides in both classical (79 ± 2%) and µ-IMER (79 ± 4%) digestions. By providing a comprehensive comparison of traditional in-gel digestion and µ-IMER methods, this study offers valuable insights into the advantages and limitations of each approach, particularly in the context of complex biological samples. The findings set a new benchmark in protein digestion and analysis, highlighting the potential of µ-IMER systems for enhanced sequence coverage and post-translational modification detection.

Metal tolerance and Cd phytoremoval ability in Pisum sativum grown in spiked nutrient solution.

In the presented study, the effects of cadmium (Cd) stress and silicon (Si) supplementation on the pea plant (Pisum sativum L.) were investigated. The tendency to accumulate cadmium in the relevant morphological parts of the plant (roots and shoots respectively)-bioaccumulation, the transfer of this element in the plant (translocation) and the physiological parameters of the plant through indicators of oxidative stress were determined. Model studies were carried out at pH values 6.0 and 5.0 plant growth conditions in the hydroponic cultivation. It was shown that Cd accumulates mostly in plant roots at both pH levels. However, the Cd content is higher in the plants grown at lower pH. The Cd translocation factor was below 1.0, which indicates that the pea is an excluder plant. The contamination of the plant growth environment with Cd causes the increased antioxidant stress by the growing parameters of the total phenolic content (TPC), polyphenol oxidase activity (PPO), the malondialdehyde (MDA) and lipid peroxidation (LP). The results obtained showed that the supplementation with Si reduces these parameters, thus lowering the oxidative stress of the plant. Moreover, supplementation with Si leads to a lower content of Cd in the roots and reduces bioaccumulation of Cd in shoots and roots of pea plants.

Screening for volatile biomarkers of colorectal cancer by analyzing breath and fecal samples using thermal desorption combined with GC-MS (TD-GC-MS).

Breath and fecal VOCs, among others, represent a new and encouraging clinical practice for the differential diagnosis of CRC. The purpose of our research was to identify VOCs present in exhaled air and feces of 20 HVs and 15 CRC patients. For collection of gas phase released from feces, emission microchambers were applied. Sorption tubes were used to enrich analytes for both breath and fecal samples. TD technique combined with GC-MS was used at the separation and identification step. The combination of statistical methods was used to evaluate the ability of VOCs to classify control group and CRC patients. Heptanoic acid, acetone, 2,6,10-trimethyldodecane, n-hexane, skatole, and dimethyl trisulfide are observed in elevated amounts in the patients group. The performance of diagnostic models on the tested data set was above 90%. This study is the first attempt to document the using of TD-GC-MS to analyze both breath and fecal samples to search for volatile biomarkers of CRC. A full evaluation of the results described herein requires further studies involving a larger number of samples. Moreover, it is particularly important to understand the metabolic pathways of substances postulated as tumor biomarkers.

A novel effective bio-originated methylene blue adsorbent: the porous biosilica from three marine diatom strains of Nanofrustulum spp. (Bacillariophyta).

In the present paper, for the first time the ability of the porous biosilica originated from three marine diatom strains of 'Nanofrustulum spp.' viz. N. wachnickianum (SZCZCH193), N. shiloi (SZCZM1342), N. cf. shiloi (SZCZP1809), to eliminate MB from aqueous solutions was investigated. The highest biomass was achieved under silicate enrichment for N. wachnickianum and N. shiloi (0.98 g L-1 DW and 0.93 g L-1 DW respectively), and under 15 °C for N. cf. shiloi (2.2 g L-1 DW). The siliceous skeletons of the strains were purified with hydrogen peroxide and characterized by SEM, EDS, the N2 adsorption/desorption, XRD, TGA, and ATR-FTIR. The porous biosilica (20 mg DW) obtained from the strains i.e. SZCZCH193, SZCZM1342, SZCZP1809, showed efficiency in 77.6%, 96.8%, and 98.1% of 14 mg L-1 MB removal under pH 7 for 180 min, and the maximum adsorption capacity was calculated as 8.39, 19.02, and 15.17 mg g-1, respectively. Additionally, it was possible to increase the MB removal efficiency in alkaline (pH = 11) conditions up to 99.08% for SZCZP1809 after 120 min. Modelling revealed that the adsorption of MB follows Pseudo-first order, Bangham's pore diffusion and Sips isotherm models.

Pyrolized Diatomaceous Biomass Doped with Epitaxially Growing Hybrid Ag/TiO2 Nanoparticles: Synthesis, Characterisation and Antibacterial Application.

In the pursuit of innovative solutions for modern technologies, particularly in the design and production of new micro/nanostructured materials, microorganisms acting as "natural microtechnologists" can serve as a valuable source of inspiration. This research focuses on harnessing the capabilities of unicellular algae (diatoms) to synthesize hybrid composites composed of AgNPs/TiO2NPs/pyrolyzed diatomaceous biomass (AgNPs/TiO2NPs/DBP). The composites were consistently fabricated through metabolic (biosynthesis) doping of diatom cells with titanium, pyrolysis of the doped diatomaceous biomass, and chemical doping of the pyrolyzed biomass with silver. To characterize the synthesized composites, their elemental and mineral composition, structure, morphology, and photoluminescent properties were analysed using techniques such as X-ray diffraction, scanning and transmission electron microscopy, and fluorescence spectroscopy. The study revealed the epitaxial growth of Ag/TiO2 nanoparticles on the surface of pyrolyzed diatom cells. The antimicrobial potential of the synthesized composites was evaluated using the minimum inhibitory concentration (MIC) method against prevalent drug-resistant microorganisms, including Staphylococcus aureus, Klebsiella pneumonia, and Escherichia coli, both from laboratory cultures and clinical isolates.

Accumulation of polystyrene nanoplastics and triclosan by a model tooth-carp fish, Aphaniops hormuzensis (Teleostei: Aphaniidae).

The presence and effects of nanoplastics (NPs; <1 µm) in the aquatic environment are a growing concern. In this study, a model tooth-carp fish, Aphaniops hormuzensis, has been exposed to different concentrations of fluorescent polystyrene nanoplastics (PS-NP) in its diet (up to 5 mg kg-1) over periods of 28 d and the particle accumulation in different tissues determined. Accumulation was observed in both digestive and non-digestive organs, with concentrations greater in the gut, liver and gill (up to 400 µg kg-1 dw) than in the skin and muscle (<180 µg kg-1 dw), but no dependency on exposure time or dose was evident. The presence of the organic contaminant, triclosan (TCS), in the diet and at concentrations up to 0.5 µg kg-1 did not affect PS-NP uptake by A. hormuzensis, while TCS accumulation in the whole body increased with time (up to 10 µg kg-1) and, likewise, appeared to be unaffected by the presence of PS-NPs. These observations suggest that the two contaminants do not interact with each other or that any interactions have no impact on accumulation. The results of this study add to the growing body of evidence that NPs can be translocated by aquatic organisms after ingestion, and reveal that, for the species and conditions employed, nanoplastics are accumulated more readily than a widely used organic chemical.

Determination of Pathogens by Electrophoretic and Spectrometric Techniques.

In modern medical diagnostics, where analytical chemistry plays a key role, fast and accurate identification of pathogens is becoming increasingly important. Infectious diseases pose a growing threat to public health due to population growth, international air travel, bacterial resistance to antibiotics, and other factors. For instance, the detection of SARS-CoV-2 in patient samples is a key tool to monitor the spread of the disease. While there are several techniques for identifying pathogens by their genetic code, most of these methods are too expensive or slow to effectively analyze clinical and environmental samples that may contain hundreds or even thousands of different microbes. Standard approaches (e.g., culture media and biochemical assays) are known to be very time- and labor-intensive. The purpose of this review paper is to highlight the problems associated with the analysis and identification of pathogens that cause many serious infections. Special attention was paid to the description of mechanisms and the explanation of the phenomena and processes occurring on the surface of pathogens as biocolloids (charge distribution). This review also highlights the importance of electromigration techniques and demonstrates their potential for pathogen pre-separation and fractionation and demonstrates the use of spectrometric methods, such as MALDI-TOF MS, for their detection and identification.

Comprehensive Study of Si-Based Compounds in Selected Plants (Pisum sativum L., Medicago sativa L., Triticum aestivum L.).

This review describes the role of silicon (Si) in plants. Methods of silicon determination and speciation are also reported. The mechanisms of Si uptake by plants, silicon fractions in the soil, and the participation of flora and fauna in the Si cycle in terrestrial ecosystems have been overviewed. Plants of Fabaceae (especially Pisum sativum L. and Medicago sativa L.) and Poaceae (particularly Triticum aestivum L.) families with different Si accumulation capabilities were taken into consideration to describe the role of Si in the alleviation of the negative effects of biotic and abiotic stresses. The article focuses on sample preparation, which includes extraction methods and analytical techniques. The methods of isolation and the characterization of the Si-based biologically active compounds from plants have been overviewed. The antimicrobial properties and cytotoxic effects of known bioactive compounds obtained from pea, alfalfa, and wheat were also described.

Fractionation of bacteria by electrophoresis as pre-separation method before MALDI-MS detection.

The present study attempted to apply the capillary electrophoresis technique for the fractionation and separation of S. Staphylococcus hominis and Escherichia coli bacteria isolated from urine samples and the detection of migrated fraction with spectrometric method. This involved the selection of suitable conditions for separation as well as the identification of pathogens. The result of the research was the separation of Gram-negative and Gram-positive bacteria, as well as their subsequent identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry using two different approaches (culture of fractions on an agar plate and direct analysis of the collected fractions). The preliminary results provide a solid basis for further research on the use of electromigration techniques with LDI detection to identify pathogens such as bacteria and viruses in biological samples.

Removal of the Basic and Diazo Dyes from Aqueous Solution by the Frustules of Halamphora cf. salinicola (Bacillariophyta).

Industrial wastes with hazardous dyes serve as a major source of water pollution, which is considered to have an enormous impact on public health. In this study, an eco-friendly adsorbent, the porous siliceous frustules extracted from the diatom species Halamphora cf. salinicola, grown under laboratory conditions, has been identified. The porous architecture and negative surface charge under a pH of 7, provided by the various functional groups via Si-O, N-H, and O-H on these surfaces, revealed by SEM, the N2 adsorption/desorption isotherm, Zeta-potential measurement, and ATR-FTIR, respectively, made the frustules an efficient mean of removal of the diazo and basic dyes from the aqueous solutions, 74.9%, 94.02%, and 99.81% against Congo Red (CR), Crystal Violet (CV), and Malachite Green (MG), respectively. The maximum adsorption capacities were calculated from isotherms, as follows: 13.04 mg g-1, 41.97 mg g-1, and 33.19 mg g-1 against CR, CV, and MG, respectively. Kinetic and isotherm models showed a higher correlation to Pore diffusion and Sips models for CR, and Pseudo-Second Order and Freundlich models for CV and MG. Therefore, the cleaned frustules of the thermal spring-originated diatom strain Halamphora cf. salinicola could be used as a novel adsorbent of a biological origin against anionic and basic dyes.

Advanced Mass Spectrometric Techniques for the Comprehensive Study of Synthesized Silicon-Based Silyl Organic Compounds: Identifying Fragmentation Pathways and Characterization.

The primary objective of this study was to synthesize and characterize novel silicon-based silyl organic compounds in order to gain a deeper understanding of their potential applications and interactions with other compounds. Four new artificial silyl organic compounds were successfully synthesized: 1-O-(Trimethylsilyl)-2,3,4,6-tetra-O-acetyl-ß-d-glucopyranose (compound 1), 1-[(1,1-dimethylehtyl)diphenylsilyl]-1H-indole (compound 2), O-tert-butyldiphenylsilyl-(3-hydroxypropyl)oleate (compound 3), and 1-O-tert-Butyldiphenylsilyl-myo-inositol (compound 4). To thoroughly characterize these synthesized compounds, a combination of advanced mass spectrometric techniques was employed, including nanoparticle-assisted laser desorption/ionization mass spectrometry (NALDI-MS), Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR-MS), and triple quadrupole electrospray tandem mass spectrometry (QqQ ESI-MS/MS). These analytical methods enabled the accurate identification and characterization of the synthesized silyl organic compounds, providing valuable insights into their properties and potential applications. Furthermore, the electrospray ionization-Fourier transform ion cyclotron resonance-tandem mass spectrometry (ESI-FT-ICR-MS/MS) technique facilitated the proposal of fragmentation pathways for the ionized silyl organic compounds, contributing to a more comprehensive understanding of their behavior during mass spectrometric analysis. These findings suggest that mass spectrometric techniques offer a highly effective means of investigating and characterizing naturally occurring silicon-based silyl organic compounds, with potential implications for advancing research in various fields and applications in different industries.

Nanostructured supports for multienzyme co-immobilization for biotechnological applications: Achievements, challenges and prospects.

The synergistic combination of current biotechnological and nanotechnological research has turned to multienzyme co-immobilization as a promising concept to design biocatalysis engineering. It has also intensified the development and deployment of multipurpose biocatalysts, for instance, multienzyme co-immobilized constructs, via biocatalysis/protein engineering to scale-up and fulfil the ever-increasing industrial demands. Considering the characteristic features of both the loaded multienzymes and nanostructure carriers, i.e., selectivity, specificity, stability, resistivity, induce activity, reaction efficacy, multi-usability, high catalytic turnover, optimal yield, ease in recovery, and cost-effectiveness, multienzyme-based green biocatalysts have become a powerful norm in biocatalysis/protein engineering sectors. In this context, the current state-of-the-art in enzyme engineering with a synergistic combination of nanotechnology, at large, and nanomaterials, in particular, are significantly contributing and providing robust tools to engineer and/or tailor enzymes to fulfil the growing catalytic and contemporary industrial needs. Considering the above critics and unique structural, physicochemical, and functional attributes, herein, we spotlight important aspects spanning across prospective nano-carriers for multienzyme co-immobilization. Further, this work comprehensively discuss the current advances in deploying multienzyme-based cascade reactions in numerous sectors, including environmental remediation and protection, drug delivery systems (DDS), biofuel cells development and energy production, bio-electroanalytical devices (biosensors), therapeutical, nutraceutical, cosmeceutical, and pharmaceutical oriented applications. In conclusion, the continuous developments in nano-assembling the multienzyme loaded co-immobilized nanostructure carriers would be a unique way that could act as a core of modern biotechnological research.

Electrothermaly conditioned carbon fibre for the analysis of volatile pollutants.

This study deals with the development of an inexpensive and single-step sorbent manufacturing methodology for the analysis of air pollutants. Disposable carbon fibre sorbents were prepared in a few minutes using the electrothermal conditioning technique. The sorbent conditioning current and time were optimised to obtain the best extraction of benzene, toluene, ethylbenzene and xylenes (BTEX) from the air samples. After sorbent characterisation, analysis parameters affecting the BTEX extraction efficiency, such as sampling volume, humidity and sampling flow rate, were optimised for active BTEX sampling. Under optimum conditions, validation parameters such as the limit of detection (LOD), repeatability, reproducibility, and linear range were found to be 0.07-0.11 mg m - 3, 1.1%-1.8%, 5.6%-9.5% and 0.24-45 mg m - 3, respectively. Thereafter, the BTEX analysis was successfully conducted using the proposed method, with acceptable recovery values (96%-103%) in the real indoor environments.

Determination of Selected Isoquinoline Alkaloids from Chelidonium majus, Mahonia aquifolium and Sanguinaria canadensis Extracts by Liquid Chromatography and Their In Vitro and In Vivo Cytotoxic Activity against Human Cancer Cells.

The search for new substances with cytotoxic activity against various cancer cells, especially cells that are very resistant to currently used chemotherapeutic agents, such as melanoma cells, is a very important scientific aspect. We investigated the cytotoxic effect of Chelidonium majus, Mahonia aquifolium and Sanguinaria canadensis extracts obtained from different parts of these plants collected at various vegetation stages on FaDu, SCC-25, MCF-7, and MDA-MB-231 cancer cells. Almost all the tested extracts showed higher cytotoxicity against these cancer cells than the anticancer drug etoposide. The highest cytotoxicity against the FaDu, SCC-25, MCF-7 and MDA-MB-231 cancer cell lines was obtained for the Sanguinaria candensis extract collected before flowering. The cytotoxicity of extracts obtained from different parts of Chelidonium majus collected at various vegetation stages was also evaluated on melanoma cells (A375, G361 and SK-MEL-3). The highest cytotoxic activity against melanoma A375 cells was observed for the Chelidonium majus root extract, with an IC50 of 12.65 µg/mL. The same extract was the most cytotoxic against SK-MEL-3 cells (IC50 = 1.93 µg/mL), while the highest cytotoxic activity against G361 cells was observed after exposure to the extract obtained from the herb of the plant. The cytotoxic activity of Chelidonium majus extracts against melanoma cells was compared with the cytotoxicity of the following anticancer drugs: etoposide, cisplatin and hydroxyurea. In most cases, the IC50 values obtained for the anticancer drugs were higher than those obtained for the Chelidonium majus extracts. The most cytotoxic extract obtained from the root of Chelidonium majus was selected for in vivo cytotoxic activity investigations using a Danio rerio larvae xenograft model. The model was applied for the first time in the in vivo investigations of the extract's anticancer potential. The application of Danio rerio larvae xenografts in cancer research is advantageous because of the transparency and ease of compound administration, the small size and the short duration and low cost of the experiments. The results obtained in the xenograft model confirmed the great effect of the investigated extract on the number of cancer cells in a living organism. Our investigations show that the investigated plant extracts exhibit very high cytotoxic activity and can be recommended for further experiments in order to additionally confirm their potential use in the treatment of various human cancers.

Determination of Some Isoquinoline Alkaloids in Extracts Obtained from Selected Plants of the Ranunculaceae, Papaveraceae and Fumarioideae Families by Liquid Chromatography and In Vitro and In Vivo Investigations of Their Cytotoxic Activity.

Alkaloids are heterocyclic bases with widespread occurrence in nature. Plants are rich and easily accessible sources of them. Most isoquinoline alkaloids have cytotoxic activity for different types of cancer, including malignant melanoma, the most aggressive type of skin cancer. The morbidity of melanoma has increased worldwide every year. For that reason, developing new candidates for anti-melanoma drugs is highly needed. The aim of this study was to investigate the alkaloid compositions of plant extracts obtained from Macleaya cordata root, stem and leaves, Pseudofumaria lutea root and herb, Lamprocapnos spectabilis root and herb, Fumaria officinalis whole plant, Thalictrum foetidum root and herb, and Meconopsis cambrica root and herb by HPLC-DAD and LC-MS/MS. For determination of cytotoxic properties, human malignant melanoma cell line A375, human Caucasian malignant melanoma cell line G-361, and human malignant melanoma cell line SK-MEL-3 were exposed in vitro to the tested plant extracts. Based on the in vitro experiments, Lamprocapnos spectabilis herb extract was selected for further, in vivo research. The toxicity of the extract obtained from Lamprocapnos spectabilis herb was tested using an animal zebrafish model in the fish embryo toxicity test (FET) for determination of the LC50 value and non-toxic doses. Determination of the influence of the investigated extract on the number of cancer cells in a living organism was performed using a zebrafish xenograft model. Determination of the contents of selected alkaloids in different plant extracts was performed using high performance liquid chromatography (HPLC) in a reverse-phase system (RP) on a Polar RP column with a mobile phase containing acetonitrile, water and ionic liquid. The presence of these alkaloids in plant extracts was confirmed by LC-MS/MS. Preliminary cytotoxic activity of all prepared plant extracts and selected alkaloid standards was examined using human skin cancer cell lines A375, G-361, and SK-MEL-3. The cytotoxicity of the investigated extract was determined in vitro by cell viability assays (MTT). For in vivo determination of investigated extract cytotoxicity, a Danio rerio larvae xenograft model was used. All investigated plant extracts in in vitro experiments exhibited high cytotoxic activity against the tested cancer cell lines. The results obtained using the Danio rerio larvae xenograft model confirmed the anticancer activity of the extract obtained from Lamprocapnos spectabilis herb. The conducted research provides a basis for future investigations of these plant extracts for potential use in the treatment of malignant melanoma.