Cadmium-affected synthesis of exopolysaccharides by rhizosphere bacteria.

AIM: Study is focused on the influence of cadmium addition to growth media on production yield, their size and molecular mass of exopolysaccharides (EPS) synthesized by three rhizosphere bacteria strains. Inhibition of bacterial growth by increasing concentrations of Cd2+ was also analysed. METHODS AND RESULTS: The highest impact of Cd2+ was noticed on the growth of Arthrobacter sp. and Rhizobium metallidurans. Chryseobacterium sp. and Arthrobacter sp. produced significantly lower when compared to R. metallidurans amounts of EPS under the influence of Cd2+ . In all bacterial strains both size and molecular mass decreased after addition of Cd2+ to growth media. It causes a change in EPS conformation to more planar, which minimizes the volume of liquid in the interglobular space next to the bacterial wall. Results confirmed strong effect of Cd2+ on the structure and synthesis of bacterial EPS what can be a key factor in the interactions between rhizosphere bacteria and host plants in heavy metal polluted soils. CONCLUSION: This work proves that due to the presence of cadmium ions, the size and conformation of EPS produced by selected bacterial strains is changed to minimize their impact on cell. We suggest that shifting in EPS conformation from bigger globular particles to the smaller planar ones could be one of the probable mechanisms of Cd resistance in metallotolerant bacteria, and finally explain increased efficiency of heavy metal phytoextraction by EPS-producing plant growth-promoting micro-organisms. SIGNIFICANCE AND IMPACT OF THE STUDY: One of the most promising remediation technique for Cd-contaminated areas is the phytoremediation in which rhizosphere bacteria play an important role by protecting plants' roots from toxic condition thus enhancing efficiency of intake. EPS secretion by bacteria is one of the most common mechanisms to protect the cell from impact of unpleasant environmental conditions, for example, toxicity of heavy metals like Cd.

Microbiology neutralization of zearalenone using Lactococcus lactis and Bifidobacterium sp.

The aim of the study was to neutralize zearalenone by lactic acid bacteria (LAB) such as Lactococcus lactis and Bifidobacterium sp. and investigate the mechanism of zearalenone (ZEA) binding. Neutralization of ZEA by LAB was confirmed by identification of binding kinetics and spectroscopic studies such as Fourier transform infrared spectroscopy (FT-IR) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The obtained results showed that the kinetic process of zearalenone binding to L. lactis is not homogeneous but is expressed with an initial rapid stage with about 90% of ZEA biosorption and with a much slower second step. In case of Bifidobacterium sp., the neutralization process is homogeneous; the main stage can be described with about 88% of ZEA biosorption. MALDI-TOF-MS measurements and FTIR analysis confirmed the uptake of zearalenone molecules by bacterial species. Moreover, the assessment of dead and live lactic acid bacteria cells after zearalenone treatment was performed using fluorescence microscopy. Graphical abstract Microbiology neutralization of zearalenone using Lactococcus lactis and Bifidobacterium sp. was confirmed by identification of binding kinetics and spectroscopic studies such as FT-IR spectroscopy and MALDI-TOF-MS spectrometry. The mechanism of ZEA binding was also investigated.

Study of silver nanoparticles synthesized by acidophilic strain of Actinobacteria isolated from the of Picea sitchensis forest soil.

AIMS: In the present work the acidophilic actinobacteria strain was used as a novel reducing agent for the cheap, green and single-step synthesis of nanostructure silver particles. Structural, morphological and optical properties of the synthesized nanoparticles have been characterized by spectroscopy, dynamic light scattering and electron microscopy approach. The antimicrobial activity of silver nanoparticles against clinical strains such as Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus mirabilis and Salmonella infantis alone and in combination with antibiotics were studied. METHODS AND RESULTS: The crystalline and stable biosynthesized silver nanoparticles ranged in size from 4 to 45 nm and were mostly spherical in shape being characterized evolving several analytical techniques. The bioAgNPs inhibited growth of most bacterial strains. The highest antimicrobial activity was observed against Ps. aeruginosa (10 mm), followed by Staph. aureus, B. subtilis and Pr. mirabilis (all 8 mm). The lower activity was noticed for E. coli and Kl. pneumoniae (6 and 2 mm, respectively). Moreover, the synergistic effect of bio(AgNPs) with various commercially available antibiotics was also evaluated. The most significant results were observed for bio(AgNPs) combined with tetracycline, kanamycin, ampicillin and neomycin, followed by streptomycin and gentamycin against E. coli, Salm. infantis and Kl. pneumoniae. The most resistant bacteria to commercial antibiotics was Pr. mirabilis. CONCLUSION: The Streptacidiphilus sp. strain CGG11n isolated from acidic soil can be used to efficiently synthesize the bioactive nanoparticles using inexpensive substances in an eco-friendly and nontoxic manner. The present work provides helpful insight into the development of new antimicrobial agents with the synergistic enhancement of the antibacterial mechanism against pathogenic micro-organisms. SIGNIFICANCE AND IMPACT OF THE STUDY: The synthesized silver bionanoparticles from Streptacidiphilus sp. strain CGG11n possess potent inhibitory effect that offers valuable contribution to pharmaceutical associations.

Study of RP HPLC Retention Behaviours in Analysis of Carotenoids.

For determination of selected carotenoids, various types of columns for high-performance liquid chromatography (HPLC) with different properties have been used. The characteristics of the laboratory-used packing material containing monomeric alkyl-bonded phases (C18, C30) and phenyl as well as phenyl-hexyl stationary phases were studied. The retention data of the examined compounds were used to determine the hydrophobicity and silanol activity of stationary phases applied in the study. The presence of the polar and carboxyl groups in the structure of the bonded ligand strongly influences the polarity of the stationary phase. Columns were compared according to methylene selectivity using a series of benzene homologues. The measurements were done using a methanol-water mobile phase. Knowledge of the properties of the applied stationary phase provided the possibility to predict the RP HPLC retention behaviours in analysis of carotenoids including lutein, lycopene and ß-carotene. The composition of the mobile phase, the addition of triethylamine and the type of stationary phase had been taken into account in designing the method of carotenoid identification. Also a monolithic column characterised by low hydrodynamic resistance, high porosity and high permeability was applied. The presented results show that the coverage density of the bonded ligands on silica gel packings and length of the linkage strongly influence the carotenoid retention behaviours. In our study, the highest retention parameters for lutein, lycopene and ß-carotene were observed for C30 and C18 stationary phase. This effect corresponds with pore size of column packing greater than 100 Å and carbon content higher than 11 %.

Effect of mobile phase pH on the retention of nucleotides on different stationary phases for high-performance liquid chromatography.

The main aim of the present investigation was to study the retention and separation of eight nucleotides with the use of seven stationary phases in RP HPLC mode. Two octadecyl columns were used; however, aminopropyl, alkylamide, cholesterol, alkyl-phosphate, and phenyl were also studied. Special attention was paid to the mobile phase buffer pH, since it appears that this factor is very influential in the case of chromatographic separation of nucleotides. The retention of nucleotides was greater for mobile phase pH 4.0 in comparison with pH 7.0 (except for octadecyl and phenyl packing). This is a consequence of protonization of polar groups present on the stationary phase surface. It was proved that aminopropyl, alkyl phosphate, alkylamide packing materials are not suitable for the resolution of nucleotides. On the other hand, cholesterol and phenyl stationary phases are alternatives for conventional octadecyl phases. Application of cholesterol packing allows separation of small, polar nucleotides, which is impossible to achieve in the case of octadecyl phase. Moreover, a phenyl support allows separation of nucleotides in a shorter time in comparison with octadecyl packing.

Linear Solvation Energy Relationships in the Determination of Specificity and Selectivity of Stationary Phases.

The retention of fifty structurally different compounds has been studied using linear solvation energy relationships. Investigations were performed with the use of six various stationary phases with two mobile phases (50/50 % v/v methanol/water and 50/50 % v/v acetonitrile/water). Packing materials were home-made and functionalized with octadecyl, alkylamide, cholesterol, alkyl-phosphate and phenyl molecules. This is the first attempt to compare all of these stationary phases synthesized on the same silica gel batch. Therefore, all of them may be compared in more complex and believable way, than it was performed earlier in former investigations. The phase properties (based on Abraham model) were used to the classification of stationary phases according to their interaction properties. The hydrophilic system properties s, a, b indicate stronger interactions between solute and mobile phase for most of the columns. Both e and v cause greater retention as a consequence of preferable interactions with stationary phase by electron pairs and cavity formation as well as hydrophobic bonds. However, alkyl-phosphate phase has different retention properties, as it was expressed by positive sign of s coefficient. It may be concluded that most important parameters influencing the retention of compounds are volume and hydrogen bond acceptor basicity. The LSER coefficients showed also the dependency on the type of organic modifier used as a mobile phase component.

New approach in determination of urinary diagnostic markers for prostate cancer by MALDI-TOF/MS.

In our study, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) is proposed as a novel tool, which can be applied to analyze lipids in urine samples. For this reason, the main aim of the study was to develop and optimize the preparation protocol for urine samples in lipidomics, using urine samples obtained from patients with diagnosed cancer and non-cancer controls. Several conditions like extraction method and types of matrices were evaluated. For this purpose, two methods for the extraction of lipids, namely modified Folch and Bligh & Dyer were employed. Furthermore, two types of matrices (α-cyano-4-hydroxycinnamic acid (HCCA) and 2,5-dihydroxybenzoic acid (DHB)) for the separation of lipids into individual components was tested. The results of this study can serve as an essential source for the selection of appropriate extraction methods and the appropriate choice of a matrix for the purification and identification of a particular class of lipid in human biological fluids. Based on it, Bligh & Dyer method associated with the usage of HCCA matrix was found to be the most effective for lipidomics using MALDI-TOF/MS. The optimized method was applied to compare the lipid profile of 139 urine samples collected from both healthy individuals and patients with prostate cancer. The tandem spectroscopic analysis allowed to identify lysophosphatidylcholine, phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and triacylglycerols in urine samples. Finally, MALDI-TOF/MS analysis enabled to discriminate between the two tested groups (healthy individuals and patients with prostate cancer). A preliminary statistical model suggested that classification accuracy ranging from 83.3 to100.0% may be achieved by using pre-selected MS signals.

Study of the Interactions of Ionic Liquids in IC by QSRR.

The nature of ionic liquids (ILs) facilitates their analysis by ion chromatography which, unlike conventional high-performance liquid chromatography, enables analysis both of cations and anions. This paper describes a pioneering ion-chromatographic investigation of IL cations and statistical evaluation of quantitative structure-retention relationships with the objective of predicting the molecular mechanism responsible for retention. Eleven ionic liquid imidazolium and pyridinium cations were analyzed on a CS15 cation-exchange column by isocratic elution with acetonitrile-methanesulfonic acid mixtures. Structural descriptors of the cations obtained from molecular modeling were used to describe their hydrophobicity as determined by chromatography. The most statistically significant were three-term QSRR regression equations relating log k(w) to analyte n-octanol-water partition coefficient (log P), dipole moment (µ), solvent accessible surface area (A(SAS)), and hydration energy (HE). They indicate the important role of both hydrophobic and polar the retention of ILs on the CS15 column.

A new method for the determination of 2,2'-dichloro-4,4'-methylenedianiline in workplace air samples by HPLC-DAD.

A new procedure has been developed for the assay of 2,2'-dichloro-4,4'-methylenedianiline (MOCA) using high-performance liquid chromatography with diode array detector. MOCA was sampled from workplace air and derivative before determination using 3,5-dinitrobenzoyl chloride. The determination was carried out in the reverse-phase system (mobile phase: acetonitrile: water) using an Ultra C(18) column. The measurement range was 2-40 µg/m(3) for a 100 dm(3) air sample. Limit of detection: 7.9 ng/m(3) and limit of quantification: 23.8 ng/m(3).

Characterization of the properties of stationary phases for liquid chromatography in aqueous mobile phases using aromatic sulphonic acids as the test compounds.

We investigated the effects of the concentration of naphthalene sulphonic acids (NSAs) as anionic test compounds in the injected sample and of the salt additives to the mobile phase on ion-exclusion. The retention behaviour of NSAs sensitively reflects even minor changes in the ionic and hydrophobic interactions and can be useful for predicting the effects of the stationary phases in reversed-phase chromatography of polar and ionic compounds, both small ones and biopolymers, e.g., oligonucleotides. We studied chromatographic properties of several stationary phases intended for separations in aqueous mobile phases: a C18 column end-capped with polar hydrophilic groups, a densely bonded C8 column doubly end-capped with short alkyl groups, a short alkyl stationary phase designed to keep full pore accessibility in highly-aqueous mobile phases and a Bidentate column with "bridged" C18 groups attached to the silica hydride support. The chemistry and pore structure of various types of column packing materials and of the salt additives to the mobile phase affect the proportion of the pore volume non-accessible to anions due to ion-exclusion and consequently the peak asymmetry and hydrophobic selectivity in reversed-phase chromatography of organic acids. We also addressed the problems connected with the determination of column hold-up volume in aqueous mobile phases. The accessibility of the stationary phase for anionic compounds in contact with the sample zone is affected by ion-exclusion due to repulsive interactions with the negatively charged surface in the pores of the stationary phase. The accessible part of the stationary phase increases and consequently the migration velocity along the column decreases with increasing concentration of the sample in the zone moving along the column. Because of a limited access to the stationary phase, its capacity can be easily overloaded. The combination of the column overload and ion-exclusion effects may result in fronting or tailing peak asymmetry. To explain this behaviour, we proposed a modified Langmuir model, respecting the variation of the column capacity due to the effects of sample concentration on ion-exclusion.

Study of Bacillus subtilis response to different forms of silver.

Although silver nanoparticles are the most widespread product of nanotechnology, the mechanisms underlying AgNP microbial toxicity remain the subject of intense debate. In this study, Bacillus subtilis has been used as model organism to elucidate the molecular interactions between this class of bacteria and different forms of silver such as nanoparticles, nanoparticles functionalized with tetracycline and silver ions. For this purpose, we carried out transmission electron microscopy and MALDI-TOF MS analysis of cells treated with silver nanoparticles (AgNPs, AgNPs functionalized with tetracycline, combination of AgNPs with tetracycline) and silver ions as well as we measured the level of reactive oxygen species. The data demonstrate that B. subtilis exhibits high resistance to silver nanoparticles and this phenomenon is associated with following processes: (I) initiation of endospore formation, (II) reduction of free Ag+ released from nanoparticles and (III) modification of the AgNPs surface. However, high silver ions concentration appeared to be very toxic to studied strain of bacterium. MALDI-TOF MS analysis revealed that the spectra of B. subtilis cells treated with silver ions are significantly different from spectra of control cells and cell treated with AgNPs and antibiotic which can suggest that silver ions in the highest degree modify bacterial components.

Zearalenone and its metabolites: Effect on human health, metabolism and neutralisation methods.

Mycotoxins are natural compounds produced as secondary metabolites by mold fungi belonging mainly to the Fusarium family, commonly found on plants such as corn or small grains in the temperate climate zone. One of these mycotoxins is zearalenone, which is classified as a xenoestrogen, an exogenous compound which resembles the structure of naturally occurring estrogens with its chemical structure. This property of zearalenone determines its ability to bind to estrogen receptors of cell and its bioaccumulation. This leads to disorders of the hormonal balance of the body, which in consequence may lead to numerous diseases of reproductive system such as prostate, ovarian, cervical or breast cancers. High risk posed by long-term exposure to contaminated food forces the modern science to develop and implement effective methods of zearalenone neutralisation. This work is a review of current state of knowledge on toxic effects of zearalenone, its metabolism in biological systems and proposed methods of its neutralisation.

Excess isotherms as a new way for characterization of the columns for reversed-phase liquid chromatography.

In this work, we present the excess isotherm of acetonitrile for stationary phases with different coverage density. Data obtained with the minor disturbance method were compared with (29)Si cross-polarization/magic-angle spinning NMR spectra to find dependence between acetonitrile adsorption on C18 chemically bonded stationary phases and coverage density of stationary phase. The preferential adsorption of acetonitrile on the bonded phase and the adsorption of water on the silica surface can be well correlated with the coverage density.

The effect of biosilver nanoparticles on different bacterial strains' metabolism reflected in their VOCs profiles.

The use of silver nanoparticles has become in recent years a growing interest for many researchers, due to their bacteriostatic and bactericidal properties and synergetic effects when they are used together with antibiotics, for an increased efficiency and less adverse reactions in the treatment of bacterial infections. Gas chromatography coupled with mass spectrometry (GC-MS), which is considered 'the golden standard' in chemical analysis, has proven to be a reliable instrument, perfectly suitable for clinical analysis. In this work, three bacterial strains, Escherichia coli (E. coli), Klebsiella oxytoca (K. oxytoca) and Staphylococcus saccharolyticus (S. saccharolyticus) were treated with biosilver nanoparticles (bioAgNPs). Headspace and GC-MS analysis was used for the detection of volatile metabolites. We observed decreased amounts of alcohols and carbonyl components (mainly ketones) in K. oxytoca and S. saccharolyticus bacteria incubated with silver. In contrast, biosilver nanoparticles added to E. coli increased the amount of VOCs, mainly hydrocarbons and alcohols. Our results have successfully demonstrated that the treatment of bacterial strains with bioAgNPs has a direct influence on their VOC profiles, by modifying the number of metabolic markers. Connected with this, the inhibition of bacteria is supposed, and consequently both the bacteriostatic and/or bactericidal effects of bioAgNPs on all three bacterial strains investigated were revealed.

Influence of clinoptilolite rock on chemical speciation of selected heavy metals in sewage sludge.

The chemical speciation of Cd, Cu, Pb, Cr and Ni in Torun municipal sewage sludge is investigated with addition of a natural sorbent (clinoptilolite rock). The total contents of the heavy metals in the sludge are substantially lower than the corresponding limits established by European or Polish legislation excepting nickel only. But the metals concentrations excepting lead exceed significantly the natural background (average contents in soils and in the Earth's crust) in dozens. Application of the sequential chemical extraction indicated that the metals in the sewage sludge are bound mainly (over 50%) in the residual fraction. The metals form the following order by parts of the mobile form: Ni> Cd>> Cr> Cu>> Pb. Addition of the clinoptilolite to the sludge leads to the metals contents fall in all four fractions of the sequential procedure. Concentrations of mobile forms of cadmium, chromium, copper and nickel decrease by 87, 64, 35 and 24%, respectively, as a result of addition of 9.09% of the clinoptilolite. The total decreases of the metals amount after 9.09% clinoptilolite addition to the sludge are around 11, 15, 25, 41 and 51% for copper, nickel, chromium, cadmium and lead, respectively.

Polymer monoliths with silver nanoparticles-cholesterol conjugate as stationary phases for capillary liquid chromatography.

Monoliths in fused-silica capillaries were prepared by an in-situ polymerization using glycidyl methacrylate (GMA) as a functional monomer and in the presence of different crosslinkers such as ethylene dimethacrylate (EDMA), trimethylolpropane trimethacrylate (TRIM), 1,6-hexanediol dimethacrylate (HEDMA), pentaerythritol tetraacrylate (PETeA), triethylene glycol dimethacrylate (TriEDMA), while azobisisobutyronitrile (AIBN) was an initiator. The monoliths' surfaces were modified chemically with cystamine dihydrochloride, followed by immobilization of Ag-NPs the surface of which was functionalized with cholesterol cysteamine (Chol-SH). Such a new material enabled the separation of proteins in a reversed phase mode. The influence of the kind of a cross-linker and composition of the porogenic solutions on the porosity and separation properties of the obtained polymers was investigated and discussed. It was found that the amount of Ag-NPs embedded on the monoliths' surfaces depended on the porous properties of the monoliths, which was directly connected with the kind of the cross-linkers used. For the determination of the amount of the attached Ag-NPs the thermogravimetric analysis was used and additionally the ISEC method as a comparative method was also proposed. It was found that the maximum amount (about 20% by weight) of the 10nm Ag-NPs was attached to the poly(GMA-EDMA) and poly(GMA-TRIM) monoliths.

Zinc oxide nanoparticles: Synthesis, antiseptic activity and toxicity mechanism.

Zinc oxide (ZnO), as a material with attractive properties, has attracted great interest worldwide, particularly owing to the implementation of the synthesis of nano-sized particles. High luminescent efficiency, a wide band gap (3.36eV), and a large exciton binding energy (60meV) has triggered intense research on the production of nanoparticles using different synthesis methods and on their future applications. ZnO nanomaterials can be used in industry as nano-optical and nano-electrical devices, in food packaging and in medicine as antimicrobial and antitumor agents. The increasing focus on nano zinc oxide resulted in the invention and development of methods of nanoparticles synthesis. Recently, various approaches including physical, chemical and biological ("green chemistry") have been used to prepare ZnO nanocomposites with different morphologies. The obtained nanoparticles can be characterized with a broad range of analytical methods including dynamic light scattering (DLS), electron microscopy (TEM, SEM), UV-VIS spectroscopy, X-ray diffraction (XRD) or inductively coupled plasma with mass spectrometry (ICP-MS). With these it is possible to obtain information concerning the size, shape and optical properties of nanoparticles. ZnO NPs exhibit attractive antimicrobial properties against bacteria (Gram-positive and Gram-negative) and fungi. Zinc oxide nanocomposites show also selective toxicity toward normal and cancerous cells, which is explained by reactive oxygen formation (ROS). Yet despite the potentially interesting antitumor activity of ZnO nanoparticles, it has been proven that they can be also cytotoxic and genotoxic for multiple types of human cells (i.e. neuronal or epithelial cells). This paper reviews the methods of synthesizing zinc oxide nanocomposites as well as their characteristics, antimicrobial activity and cytotoxicity against normal and tumor cells.

Monolithic continuous beds as a new generation of stationary phase for chromatographic and electro-driven separations.

Monolithic supports are a new generation of stationary phases which are typically prepared using a simple molding process carried out within the confines of the capillary. They provide high rates of mass transfer at lower pressure drops, enable much faster separations and the nature of the pores allows easy permeability for large molecules. This review summarizes the current achievements and application of organic polymer-based monolithic columns, silica-based monolithic columns and their application in bioaffinity processing, modern biotechnology, determination of microorganisms and chiral separations. Special attention is paid to microfabricated devices with monolithic supports because their fabrication of particles directly in the channel eliminates the need for a frit and also creates a unique homogeneity of packing.

Hypercrosslinked cholesterol-based polystyrene monolithic capillary columns.

Cholesteryl methacrylate has been used to obtain hypercrosslinked poly(cholesteryl methacrylate-co-4-methylstyrene-co-vinylbenzyl chloride-co-divinylbenzene) monolithic stationary phase for capillary liquid chromatography. Different contents of cholesteryl methacrylate and toluene in polymerization mixture have been tested to obtain chromatographic performance. The best column has been prepared using 7% of cholesteryl methacrylate and 1% of toluene in polymerization mixture. Such a monolithic material exhibited a surface area of 452m2/g (BET). The column efficiency of about 63 000 plates/m (H=15µm) was achieved for benzene. The polystyrene column with cholesteryl methacrylate providing an additional functional group gave the highest efficiency compared to known poly(4-methylstyrene-co-vinylbenzyl chloride-co-divinylbenzene) and poly(cholesteryl methacrylate-co-trimethylolpropane trimethacrylate) monolithic columns. All of the above capillary columns were used for the separations of small molecules, such as alkylbenzenes, o-terphenyl/triphenylene, parabens, and proteins in reversed-phase liquid chromatography mode. The relative standard deviation of day-to-day reproducibility of hypercrosslinked poly(cholesteryl methacrylate-co-4-methylstyrene-co-vinylbenzyl chloride-co-divinylbenzene) monolithic columns were between 12.39-15.95% for retention factors and 10-24% for plate heights, respectively.

Determination of some psychotropic drugs in serum and saliva samples by HPLC-DAD and HPLC MS.

A simple, rapid and sensitive HPLC-DAD method has been developed and validated for the simultaneous determination of seven psychotropic drugs (risperidone, citalopram, clozapine,quetiapine, levomepromazine, perazine and aripiprazole) in human serum or saliva samples. The chromatographic analyses were performed on a XSELECT CSH Phenyl-Hexyl column with a mobile phase containing methanol, acetate buffer at pH 3.5 and 0.025mL(-1) diethylamine. The influence of concentration of methanol in injection samples and injection volume on peak symmetry and system efficiency was examined.The full separation of all investigated drugs, good peaks' symmetry and simultaneously high systems efficiency were obtained in applied chromatographic system. The method is suitable for the analysis of investigated drugs in human plasma or saliva for psychiatric patients for control of pharmacotherapy, particularly in combination therapy. HPLC-MS was applied for verification of the presence of drugs and their metabolites in serum and saliva samples from patients.