Combination Of High-performance Liquid Chromatography Research Articles

Trinitroaromatic explosives: Modern application, toxicological characterization, and methods of determination

Explosives such as tetryl and picric acid, which were common in the past, now have lost their combat relevance. However, they are often used for peaceful purposes individually and in combination with other trinitroaromatic compounds (e.g., trinitrotoluene). As a result of their use, environmental pollution occurs, followed by intoxication of plants, animals, and people. Cases of explosive poisoning during their production are also described.
 The symptoms of poisoning include both of general disorders and specific phenomena such as skin staining, impaired physiological efficiency of NADPh-dependent enzymes, genotoxicity, and immunotoxicity.
 Previous scientific studies established a trend toward the development of chemical-analytical probes. Various options for the sensor surface of the device and methods for detecting compounds are considered. To determine the explosives, ion mobility spectrometry is widely used, which is very rare for the chemicaltoxicological analysis of other groups of compounds.
 Simultaneously, methods commonly used in the analysis of narcotic and psychotropic substances (gas chromatography/ combination of high-performance liquid chromatography and mass spectrometry methods) are also applicable to determine trinitroaromatic explosives. However, the presence of nitro groups in their structure complicates such an analysis. This problem can be resolved by injecting cold samples directly to the column.
 Despite the availability of various developed techniques and methods, the possibility of their application to study biological matrices remains insufficient.
 Therefore, further studies of the chemicaltoxicological nature should be conducted to establish the optimal conditions for extracting the substances in question, the parameters of instrumental analysis, and the possibility of storing samples and for solving other problems of forensic medical examination.

Quality Control and Authentication of Black Betel Leaf Extract (Piper acre Blume) from East Kalimantan as an Antimicrobial Agent Using a Combination of High-Performance Liquid Chromatography and Chemometric Fourier Transform Infrared.

Black betel leaf from East Kalimantan contains various secondary metabolites such as alkaloid saponins, flavonoids, and tannins. A compound, piperenamide A, which has antimicrobial activity, is also found in black betel leaf. This study aims to identify and authenticate the compound piperenamide A found in black betel leaf extract in other types of betel plant using HPLC and FTIR-chemometrics. The extraction method used was maceration with 70% ethanol solvent. Determination of piperenamide A content in black betel leaf extract was via HPLC column C18, with a maximum wavelength of 259 nm and a mobile phase of water:acetonitrile at a flow rate of 1 mL/minute. From the results, piperenamide A was only found in black betel (Piper acre) and not in Piper betel and Piper crocatum. Piperenamide A levels obtained were 4.03, 6.84, 5.35, 13.85, and 2.15%, respectively, in the samples studied. The combination of FTIR spectra with chemometric methods such as PCA and PLS-DA was used to distinguish the three types of betel. Discriminant analysis can classify black betel (Piper acre), Piper betel, and Piper crocatum according to its type. These methods can be used for identification and authentication of black betel.

Secondary metabolites of Schisandra chinensis in homeostasis regulator adaptogen herbal formula for preventive oncology

The original herbal formula of homeostasis regulator Multiphytoadaptogen (MPhA) for preventive oncology developed by the N.N. Blokhin Center of Oncology containing phytocomponents from Schizandra chinensis has been investigated in vitro, in vivo and in clinical studies. The MPhA multi-target effects are achieved by optimizing the functioning of the nervous, immune and endocrine defense systems that regulate homeostasis under stress. Everything that has been previously studied for MPhA can be considered as preclinical testing, including clinical research, which can be regarded as the pilot studies. This was allowed because MPhA in Russia is registered as a parapharmaceutical agent and therefore standardized according to established requirements. However, due to the high efficiency of MPhA, a detailed study of the chemical composition and standardization of it is required, including the composition of Schisandra chinensis Baill (Schisandraceae) active components, which turned out to be translocated into MPhA as a result of the extraction technology developed. So, for MPhA identification and standardization we detected the secondary metabolites in the herbal formula MPhA as well as in fruits extract of Schisándra chinénsis using high-performance liquid chromatography in combination with mass spectrometry. Chromatography was performed on an ACQUITY UPLC BEH C18 column in a gradient mode. A TSQ Vantage triple quadrupole mass spectrometer with electrospray ionization was used. Lignans Schizandrin and Schizantherin A were identified in the MPhA as well as in Schisándra chinénsis fruits extract obtained by the technology developed. The determined secondary metabolites can be used for standardization and quality testing of the herbal formula MPhA. In addition, we performed in silico analyzes of Schizandrin and Schizantherin A biological activity spectra using computer program PASS (Prediction of Activity Spectra for Substances). Schizandrin and Schizantherin A activities, according the scientific literature and in silico analysis, correspond to the properties studied for MPhA which therefore fits into the concept of a drug – homeostasis regulator adaptogen for preventive oncology.

Green Extraction Techniques for the Determination of Active Ingredients in Tea: Current State, Challenges, and Future Perspectives

In recent years, the scientific community has turned its attention to the further study and application of green chemistry as well as to sustainable development in reducing the consumption of raw materials, solvents, and energy. The application of green chemistry aims to ensure the protection of the environment and to also, consequently, improve the quality of human life. It offers several benefits, both socially and economically. In the last few decades, new alternative non-conventional green extraction methodologies have been developed for the purposes of the extraction of active ingredient compounds from various raw products. The main objective of this literature review is to present the current knowledge and future perspectives regarding the green extraction of tea species in respect of the isolation of safe active biomolecules, which can be used as commercially available products—both as dietary supplements and pharmaceutical formulations. More specifically, in this literature review, the intention is to investigate several different extraction techniques, such as ultrasonic-assisted extraction, ultrasonic-assisted extraction with DESs, the microwave assisted-extraction method, and the reflux method. These are presented in respect of their role in the isolation of bioactive molecules regarding different tea species. Furthermore, following the literature review conducted in this study, the commonly used green extraction methods were found to be the ultrasound-assisted method and the microwave-assisted method. In addition to these, the use of a green solvent, in regard to its role in the maximum extraction yield of active ingredients in various species of tea, was emphasized. Catechins, alkaloids (such as caffeine), gallic acid, and flavonoids were the main extracted bioactive molecules that were isolated from the several tea species. From this literature review, it can be demonstrated that green tea has been widely studied at a rate of 52% in respect of the included research studies, followed by black tea at 26%, as well as white tea and oolong tea at 11% each. Regarding the determination of the bioactive molecules, the most utilized analytical method was found in the combination of high-performance liquid chromatography (HPLC) with a photodiode array detector (PDA) and mass spectrophotometry (MS) at a usage rate of about 80%. This method was followed by the utilization of UPLC and GC at 12% and 8%, respectively. In the future, it will be necessary to study the combination of green extraction techniques with other industry strategies, such as an encapsulation at the micro and nano scale, for the purposes of preparing stable final products with antioxidant properties where, finally, they can be safely consumed by humans.

Comparative analysis of tryptophan and downstream metabolites of the kynurenine and serotonin pathways in patients with arterial hypertension and coronary artery disease

Aim To compare serum concentrations of tryptophane (Trp) and its metabolites in subjects with no cardiovascular disease (CVD) and patients with СVD, including arterial hypertension (AH) and ischemic heart disease (IHD).Material and methods This study included 131 participants; 58 participants (11 of them with documented peripheral atherosclerosis) were included into the AH group, 46 participants were included into the IHD group, and 27 participants with no signs of CVD were included into the control group. Plasma concentrations of Trp and its metabolites were measured by high-performance liquid chromatography in combination with a triple quadrupole analyzer.Results Comparison of the three study groups revealed significant differences in concentrations of Trp (р=0.029), kynurenine (p<0.001), kynurenine/Trp ratio (p<0.001), quinolinic acid (р=0.007), kynurenic acid (р=0.003), serotonin (p<0.001), and 5‑hydroxyindoleacetic acid (5‑HIAA) (р=0.011). When the AH group was subdivided into subgroups without and with documented peripheral atherosclerosis, the intergroup differences remained for concentrations of kynurenine, kynurenine/Trp ratio, quinolinic acid, kynurenic acid, serotonin, and 5‑HIAA. Also, correlations were found between concentrations of Trp metabolites and laboratory and instrumental data, primarily inflammatory markers.Conclusion Analysis of serum concentrations of Trp and its metabolites in CVD patients showed increases in kynurenine, kynurenine/Trp ratio, quinolinic acid, kynurenic acid, and 5‑HIAA along with decreases in concentrations of Trp and serotonin in the groups of AH, AH with documented peripheral atherosclerosis, and IHD.

Development and validation of a method for airborne monoclonal antibodies to quantify workplace exposure

Modern therapy strategies are based on patient-specific treatment where the drug and dose are optimally adapted to the patient's needs. In recent drugs, monoclonal antibodies (mAbs) are increasingly used as active ingredients. Their patient-specific formulations are not part of the pharmaceutical industry’s manufacturing process but are prepared from concentrates by pharmaceutical personnel. During the manufacturing process, however, active pharmaceutical ingredients are released in trace amounts or, in the case of accidents and spills, also in high concentrations. Regardless of the source of entry, mAbs can become airborne, be inhaled, and cause undesirable side-effects such as sensitization. To assess the risk for pharmaceutical personnel, a personal air sampling method was developed and validated for bevacizumab, cetuximab, daratumumab, omalizumab, rituximab and trastuzumab. The method is based on the combination of high-performance liquid chromatography with tandem mass spectrometry (HPLC-MS/MS). The analytical method achieves a limit of detection of 0.30–8.8 ng mL−1, recoveries of 83–96 % (intra-day assay) and 75–89 % (inter-day assay), with no detectable carry-over. A polycarbonate filter proved suitable for sampling airborne monoclonal antibodies, as it achieved 80–104 % recovery across all mAbs. It also showed concentration-independent desorption efficiency. The sampling duration can be up to 480 min without negatively affecting the recovery. MAbs are stable on the polycarbonate filter at 5 °C for 3 days (recovery: 94 % ± 5 %) and at − 20 °C for 14 days (recovery: 97 % ± 4 %). Our method demonstrated that there is a potential for release when handling monoclonal antibodies. However, this can be reduced below the limit of detection by using pressure equalization systems (spikes).

Chromium speciation by the combination of high-performance liquid chromatography and inductively coupled plasma-optical emission spectrometry.

A hyphenated instrumental method high-performance liquid chromatography-inductively coupled plasma-optical emission spectrometry (HPLC-ICP-OES) was used for the separation and determination of Cr(III) and Cr(VI). After the chromatographic separation of chromium species by anion exchange column, their spectrophotometric detection was carried out by ICP-OES system. Important instrumental and chromatographic parameters were investigated via univariate optimization approach to obtain high signal to noise ratio and good resolution for chromium species. Under the optimum HPLC-ICP-OES conditions, limit of detection (LOD) values for Cr(III) and Cr(VI) were found to be 0.27 and 0.05mg/kg, respectively. In addition, accuracy and applicability of developed method were checked by recovery experiments performed with the spiked soil, grass, and water samples. High percent recovery results (88-104%) were achieved by utilizing matrix matching calibration strategy for the selected samples. The results showed that this method was accurate and applicable to soil, grass, and water samples.

Development of a Novel HPLC-MS Method to Separate Polar and Non-Polar Compounds in Biodiesel/Petrodiesel Mixtures

Due to a trend to higher sustainability, biodiesel is often mixed into petrodiesel. The analysis of these blends on a molecular level is not trivial, since huge differences in concentrations and polarity of the analytes require a large dynamic range of the analytical method, as well as the ability to investigate molecules of widely different polarities. A combination of high-performance liquid chromatography (HPLC) with high resolution mass spectrometry (HRMS) was identified as a promising method and a normal-phase (NP)-HPLC using amino-functionalized silica gel-based stationary phase delivered the best results with very fast (under 4 min) measurements, with distinct separation of the compounds and clean mass spectra of singular compounds. This method can also be easily modified to elute all FAMEs (fatty acid methyl esters) in one singular peak, thus making the separation even faster (under 3 min).

Biochar nanosphere- and covalent organic framework nanosphere-functionalized titanium dioxide nanorod arrays on carbon fibers for solid-phase microextraction of organic pollutants

• Biochar nanospheres were grown in situ onto TiO 2 nanorod arrays to improve the extraction effect. • COF nanospheres were bonded to TiO 2 nanorod arrays to improve the extraction performance. • The structure-performance relationships and extraction mechanisms of extraction materials were summarized. • Three online IT-SPME-HPLC analytical methods were developed to detect organic pollutants. • These materials exhibited better extraction performance than some other materials. To improve the extraction efficiency of carbon fibers (CFs) toward organic pollutants, TiO 2 nanorod arrays (NARs) were grown in situ on CFs. Subsequently, biochar nanospheres and covalent organic framework nanospheres were separately introduced to functionalize the NARs. A sequence of materials was produced by regulating the reactant concentrations and characterized using a scanning electron microscope, an X-ray photoelectron spectrometer, an X-ray diffractometer, a Raman spectrometer, and a specific surface area analyzer. The materials were then deposited into separate poly(etheretherketone) tubes for in-tube solid-phase microextraction (IT-SPME). These tubes were evaluated with different types of organic pollutants (polycyclic aromatic hydrocarbons (PAHs), estrogens, bisphenols, and phthalate esters) using a combination of high-performance liquid chromatography and IT-SPME, and they exhibited diverse extraction performance. The extraction mechanism of each material is carefully discussed, and the structure–performance relationship is also summarized based on the chemical structures and extraction properties of the materials. The most efficient extraction materials for different analytes were discovered and used to develop analytical methods. Three online methods were used to sensitively detect PAHs, estrogens, and bisphenols in real water samples, respectively. Satisfactory results were obtained, including enrichment factors up to 6784, detection limits as low as 0.001 μg L −1 , linear ranges of 0.003–15.0 μg L −1 , and relative standard deviations ranging from 0.2% to 15.2%. The results indicate that these methods have some advantages over previous material-based methods.

Spatial Distribution of Polyphenolic Compounds in Corn Grains (Zea mays L. var. Pioneer) Studied by Laser Confocal Microscopy and High-Resolution Mass Spectrometry.

Desirable changes in the biochemical composition of food plants is a key outcome of breeding strategies. The subsequent localization of nutritional phytochemicals in plant tissues gives important information regarding the extent of their synthesis across a tissue. We performed a detailed metabolomic analysis of phytochemical substances of grains from Zea mays L. (var. Pioneer) by tandem mass spectrometry and localization by confocal microscopy. We found that anthocyanins are located mainly in the aleurone layer of the grain. High-performance liquid chromatography in combination with ion trap tandem mass spectrometry revealed the presence of 56 compounds, including 30 polyphenols. This method allows for effective and rapid analysis of anthocyanins by plotting their distribution in seeds and grains of different plants. This approach will permit a more efficient screening of phenotypic varieties during food plant breeding.

Isolation and structure elucidation of caryophyllane sesquiterpenoids from leaves of Eremophila spathulata

• This is the first phytochemical investigation of the leaves of Eremophila spathulata . • A combination of HPLC and HPLC-PDA-HRMS-SPE-NMR afforded seven compounds. • The structures were identified as new caryophyllane sesquiterpenoids by HRMS and 2D NMR. Crude extract of Eremophila spathulata leaves was investigated by semi-preparative scale high-performance liquid chromatography (HPLC), analytical scale HPLC, and hyphenated high-performance liquid chromatography-photodiode array-high-resolution mass spectrometry-nuclear magnetic resonance (HPLC-PDA-HRMS-SPE-NMR), which afforded seven previously unreported caryophyllane sesquiterpenoids. Semi-preparative scale separation of the crude extract afforded (1 R* ,4 R *,9 S* , E )-8-formyl-11,11-dimethylbicyclo[7.2.0]undec-7-ene-4-carboxylic acid ( 5 ) and analytical-scale HPLC separation afforded (1 R* ,4 S* ,7 S *,9 S* )-7-hydroxy-11,11-dimethyl-8-methylenebicyclo[7.2.0]undecane-4-carboxylic acid ( 1 ), (1 S* ,6 R* ,9 R*,E )-10,10-dimethylbicyclo[7.2.0]undec-2-ene-2,6-dicarboxylic acid ( 2 ), (1 R* ,4 S* ,9 S* )-11,11-dimethyl-8-oxobicyclo[7.2.0]undecane-4-carboxylic acid ( 3 ), and (1 R* ,4 R* ,9 S* )-11,11-dimethyl-8-oxobicyclo[7.2.0]undecane-4-carboxylic acid ( 4 ). HPLC-PDA-HRMS-SPE-NMR afforded (1 R* ,4 R* ,9 S* )-11,11-dimethyl-8-methylenebicyclo[7.2.0]undecane-4-carboxylic acid ( 6 ) and (1 R* ,4 S* ,9 S* )-11,11-dimethyl-8-methylenebicyclo[7.2.0]undecane-4-carboxylic acid ( 7 ). The structures of all isolated compounds were established based on HRMS as well as extensive 1D and 2D NMR analysis. Relative configurations were determined by correlations in spectra from rotational Overhauser effect spectroscopy.

Compound traditional Chinese medicine dermatitis ointment ameliorates inflammatory responses and dysregulation of itch-related molecules in atopic dermatitis

BackgroundAtopic dermatitis (AD) is a chronic inflammatory skin disease accompanied with itchy and scaly rash. Compound traditional Chinese medicine dermatitis ointment (CTCMDO) consists of a mixture of extracts from five plants, which had been used in AD treatment due to good anti-inflammatory and anti-allergic effects.Materials and methodsIn this study, high-performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometer (LC/MS) were performed to analyze the active ingredients of CTCMDO in detail and to establish its HPLC fingerprint. Furthermore, the anti-inflammatory and antipruritic activities of CTCMDO were studied in the treatment of DNCB-induced AD in mice.ResultsA total of 44 compounds including phenylpropionic acid compounds, alkaloid compounds, curcumin compounds and lignans were identified via combined HPLC and LC/MS. A fingerprint with 17 common peaks was established. In AD-like mice, DNCB-induced scratching behavior had been suppressed in the treatment of CTCMDO in a dose-dependent manner. Furthermore, the detailed experimental results indicated that the AD can be effectively improved via inhibiting the production of Th1/2 cytokines in serum, reversing the upregulation of substance P levels of itch-related genes in the skin, and suppressing the phosphorylation of JNK, ERK, and p38 in the skin.ConclusionThis work indicated that CTCMDO can significantly improve AD via attenuating the pathological alterations of Th1/2 cytokines and itch-related mediators, as well as inhibiting the phosphorylation of mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB).

Diagnostic algorithm of the subclinical forms of adrenal neoplasms

The possibility of modifying the existing diagnostic algorithm of latent forms of hormone-active and potentially malignant formations in the adrenal glands using modern methods of laboratory diagnostics and radiation imaging is substantiated. In the clinic of a faculty of surgery and on its clinical base, experience in the examination and treatment of 1457 patients with various formations of the adrenal glands has been accumulated. Among them, 270 (14.9%) patients were selected, in whom precursor hormones of steroidogenesis and metabolites of catecholamines in the blood plasma were specially studied by high-performance liquid chromatography, tumor marker measurement, and three-phase computed tomography with intravenous contrast enhancement. Determination of steroidogenesis precursor hormones and catecholamine metabolites in the blood plasma by high-performance liquid chromatography in combination with traditional methods of laboratory diagnostics, use of computed tomography with intravenous contrast enhancement, construction of multiplanar reconstructions, and postprocessor image processing are the basis of a modified algorithm for verification of latent forms of hormone-active and potentially malignant tumors of the adrenal glands. The implementation of the diagnostic algorithm developed in the clinic made early diagnosis of the subclinical forms of NP neoplasms possible, as well as ensure the implementation of minimally invasive surgical interventions before the development of endocrine and metabolic disorders and consequently prevent the development of intraoperative and postoperative complications, which thereby reduce the duration of rehabilitation and disability of patients with good long-term treatment results and high quality of life.

Identification of β1-3 galactosylglucose-core free-glycans in human urine

We previously reported the precise structure of acidic free-glycans in human urine. In the present study, structural analysis of neutral free-glycans in urine was performed in fine detail. Urine samples were collected from 21 healthy volunteers and free-glycans extracted from the creatinine-adjusted urine and then fluorescently labeled with 2-aminopyridine. Neutral glycan profiling was achieved by a combination of high-performance liquid chromatography, mass spectrometry, enzymatic digestion, and periodate cleavage. A total of 79 glycans were identified. Because the ABO-blood group antigen containing urinary neutral glycans are major components, profiling patterns were similar between individuals of the same ABO-group. The neutral glycans were composed of lactose-core (Galβ1-4Glc) glycans, type-II N-acetyllactosamine-core (GlcNAcβ1-4Glc) glycans, hexose oligomers, N-glycans and to our surprise β1-3 galactosylglucose-core (Galβ1-3Glc) glycans. Although glycans with a β1-3 galactosylglucose-core were identified as major components in urine, comprising structurally simple isomers of a lactose-core, the core structure has not previously been reported. The major β1-3 galactosylglucose-core glycans were Fucα1-2Galβ1-3(Fucα1-4)Glc, GalNAcα1-3(Fucα1-2)Galβ1-3(Fucα1-4)Glc and Galα1-3(Fucα1-2)Galβ1-3(Fucα1-4)Glc, corresponding to H-, A-, and B-blood group antigens, respectively. Three lactosamine extended β1-3 galactosylglucose-core glycans were also detected as minor components. Elucidating the biosynthesis of β1-3 galactosylglucose will be crucial for understanding the in vivo function of these glycans.

Methods for the Preparation of Blood Serum Samples in the Assessment of the Pharmacokinetics of Protein Drugs by the Example of Viscumin

A comparative evaluation of methods for preparing blood serum samples for determination of the lectin content of the mistletoe Viscum Album, viscumin, by high-performance liquid chromatography in combination with high-resolution mass-spectrometry detection has been carried out. Based on the results of the analysis of the lectin hydrolyzate, a specific peptide fragment with m/z 791.388 suitable for subsequent quantitative determination was selected. Isolation of viscumin from serum components was carried out without the use of specific antibodies. The study used methods employing various spin columns and the method of protein precipitation with 1% TCA (trichloroacetic acid) in IPA (isopropyl alcohol). Testing the methods of depletion of blood serum proteins was based on the determination of the degree of extraction of lectin from model serum samples. As a result of our research, we have developed a technique for the quantitative analysis of viscumin in blood serum by the most efficient method of sample preparation using the ProteoMiner spin columns. The technique allows to determine the protein concentration up to 5·10-4 mg/ml with the extraction degree (2.7 ± 0.6) %.

Biological functions of cobalt and its toxicology and detection in anti-doping control

Objectives. Over the last decade, hematopoietic stimulants have grown increasingly popular in elite sports. This is supported by the growing number of high-profile doping scandals linked to their use. A group of these stimulants includes cobalt salts, which cause an increase in the oxygen capacity of the blood as well as a powerful stimulation of metabolic processes, resulting innoticeable competitive advantages. The use of cobalt salts is regulated according to the Prohibited List of the World Anti-Doping Agency (WADA). Currently, only a few works have been dedicated to solving the problem of detecting the abuse of cobalt salts in anti-doping control. Only a few laboratories have included cobalt salt determination in their methodological bases. The purpose of this review is to attract the attention of the scientific community to the toxicity of cobalt compounds, consequences of their intake, and pharmacokinetics, as well as the problems in their detection methods due to their widespread availability in the modern market and the growing number of abuse cases.Results. The main biological functions of cobalt, cellular levels of exposure, toxicity, and symptoms of cobalt salt poisoning are presented in detail in this review article. The data from the literature on the main methods for detecting cobalt as a doping agent have been generalized and systematized. There is a major focus on the amount of cobalt in dietary supplements that could cause an athlete to test positive for cobalt when they are consumed.Conclusions. After analyzing promising cobalt detection approaches and methods, it was determined that high-performance liquid chromatography in combination with inductively coupled plasma mass spectrometry has an undeniable advantage for detecting cobalt as a doping agent. The lack of explicit WADA requirements for detection methods and the lack of its obligation to determine cobalt make it tempting for unscrupulous athletes to use its salts. Therefore, antidoping laboratories must implement the abovementioned method as soon as possible.

Multiple Chromatographic Analysis of Urine in the Detection of Bladder Cancer

Bladder cancer (BC) is the most common type of carcinoma of the urological system. Recently, there has been an increasing interest in non-invasive diagnostic tumor markers due to the invasive attribute of cystoscopy, which is still considered the gold standard diagnostic method. However, markers published in the literature so far do not meet expectations for replacing cystoscopy due to their low specificity and excessively high false-positive results, which can be mainly caused by frequently occurring hematuria also in benign cases. No reliable non-invasive method has yet been identified that can distinguish patients with bladder cancer and non-malignant hematuria patients. Our work examined the possibilities of non-targeted biomarkers of urine to distinguish patients with malignant and non-malignant diseases of the bladder using 3D HPLC in combination with computer processing of multiple datasets. Urine samples from 47 patients, 23 patients with bladder cancer (BC) and 24 patients with non-malignant hematuria (NMHU), were enrolled in clinical trials. For the separation and subsequent analysis of a large number of urine components, 3D HPLC (high-performance liquid chromatography) with an absorption and fluorescence detector was used. The obtained dataset was further subjected to various uni- and multi-dimensional statistical analyses and mathematical modeling. We found 334 chromatographic peaks, of which 18 peaks were identified as significantly different for BC and NMHU patients. Using receiver operating characteristic (ROC) analysis, we assessed the informative ability of significant chromatographic peaks (90% sensitivity and 74% specificity). By logistic regression, we identified the optimal and simplified set of seven chromatographic peaks (5 absorptions plus 2 fluorescence) with strong classification power (100% sensitivity and 100% specificity) for distinguishing patients with bladder cancer and those with non-malignant hematuria. Partial least square discriminant analysis (PLS-DA) model and orthogonal projection to latent structure discriminant analysis (OPLS-DA) with 100% sensitivity and 96% specificity were used to distinguish BC and NMHU patients. Multivariate statistical analysis of urinary metabolomic profiles of patients revealed that BC patients can be discriminated from NMHU patients and the results can likely contribute to an early and non-invasive diagnosis of BC.

Antiviral drug Umifenovir (Arbidol) in municipal wastewater during the COVID-19 pandemic: Estimated levels and transformation

An indole derivative umifenovir (Arbidol) is one of the most widely used antiviral drugs for the prevention and treatment of COVID-19 and some other viral infections. The purpose of the present study was to shed light on the transformation processes of umifenovir in municipal wastewater, including disinfection with active chlorine, as well as to assess the levels of the antiviral drug and its metabolites entering and accumulating in natural reservoirs under conditions of the SARS-CoV-2 pandemic. The combination of high-performance liquid chromatography with electrospray ionization high-resolution mass-spectrometry and inductively coupled plasma mass spectrometry was used for tentative identification and quantification of umifenovir and its transformation products in model reaction mixtures and real samples of wastewater, river water, biological sludge and bottom sediments taken at the wastewater treatment plant in Arkhangelsk, a large cultural and industrial center at the Russian North. Laboratory experiments allowed identifying fifteen bromine-containing transformation products, forming at the initial stages of the chlorination and fourteen classic volatile and semi volatile disinfection by-products with bromoform as the dominant one. Chlorinated derivatives are only the minor disinfection by-products forming by substitution of alkylamine group in the aromatic ring. The schemes of umifenovir transformation in reactions with dissolved oxygen and sodium hypochlorite are proposed. Two established primary transformation products formed by oxidation of the thioether group to sulfoxide and elimination of thiophenol were detected in noticeable concentrations in the wastewater together with their precursor. The level of umifenovir reached 1.3 mg kg−1 in the sludge and municipal wastewater treat contained 1 μg L−1 of that drug, while its removal during biological wastewater treatment was about 40%. Pronounced accumulation of umifenovir and its transformation products in biological sludge and bottom sediments of natural reservoirs may be a source of the future secondary pollution of the environment.

Discovery of eight alkaloids with D1 and D2 antagonist activity in leaves of Nelumbo nucifera Gaertn. Using FLIPR assays

Ethnopharmacological relevanceDopamine receptors are long-standing primary targets in the treatment of mental diseases and there is growing evidence that suggests relationships between obesity and the dopamine system, especially dopamine D1 and D2 receptors. Leaves of Nelumbo nucifera Gaertn. (lotus leaves) have been medically used for helping long-term maintenance of weight loss. Whether and how components of lotus leaves function through the dopamine receptors remains unclear. Aim of the studyThis work aimed to discover dopamine receptor-active alkaloids isolated from the lotus leaves, to evaluate their potencies and to analyze their structure activity relationship. Materials and methodsDried lotus leaves were prepared and total extract was divided into alkaloids and flavones. Eight alkaloids were separated and characterized by a combination of high-performance liquid chromatography, quadrupole time-of-flight mass spectrometry and nuclear magnetic resonance, and assayed by a fluorometric imaging plate reader platform. Human embryonic kidney 239 cell lines expressing dopamine D1, D2 and serotonin 2A (5-HT2A) receptors, respectively, were cultured and used in the assay. ResultsAlkaloids in the lotus leaves were the bioactive phytochemicals and inhibited dopamine from accessing the D1 and D2 receptors. All eight compounds functioned as D1-receptor antagonists and except N-nornuciferine, seven alkaloids functioned as D2-receptor antagonists. (S)-coclaurine and (R)-coclaurine are optical isomers and antagonized both D1 and D2 with equivalent potencies, suggesting that the optical rotation of the methylene linker in the monobenzyl isoquinoline backbone did not influence their activity. Among the eight alkaloids, O-nornuciferine was the potent antagonist to both receptors (the lowest IC50 values, D1: 2.09 ± 0.65 μM and D2: 1.14 ± 0.10 μM) while N-nornuciferine was found to be the least potent as it moderately antagonized D1 and was inactive on D2. O-nornuciferine was also a 5-HT2A antagonist (IC50~20 μM) while N-nornuciferine had no activity. These hinted the importance of a methyl group attached to the nitrogen atom in the aporphine backbone. Armepavine showed a nearly 10-fold selectivity to D2. ConclusionsIn this work, eight alkaloids were isolated from the leaves of Nelumbo nucifera Gaertn. and assayed on the D1 and D2 receptors. They were D1/D2 antagonists with IC50 values in the mid- to low-micromolar range and O-nornuciferine was the most potent alkaloid among the eight. This family of alkaloids was biochemically evaluated on the dopamine receptors by the same platform for the first time.

Plasma Oxylipins: A Potential Risk Assessment Tool in Atherosclerotic Coronary Artery Disease.

Background: While oxylipins have been linked to coronary artery disease (CAD), little is known about their diagnostic and prognostic potential.Objective: We tested whether plasma concentration of specific oxylipins may discriminate among number of diseased coronary arteries and predict median 5-year outcomes in symptomatic adults.Methods: Using a combination of high-performance liquid chromatography (HPLC) and quantitative tandem mass spectrometry, we conducted a targeted analysis of 39 oxylipins in plasma samples of 23 asymptomatic adults with low CAD risk and 74 symptomatic adults (≥70% stenosis), aged 38–87 from the Greater Portland, Oregon area. Concentrations of 22 oxylipins were above the lower limit of quantification in >98% of adults and were compared, individually and in groups based on precursors and biosynthetic pathways, in symptomatic adults to number of diseased coronary arteries [(1) n = 31; (2) n = 23; (3) n = 20], and outcomes during a median 5-year follow-up (no surgery: n = 7; coronary stent placement: n = 24; coronary artery bypass graft surgery: n = 26; death: n = 7).Results: Plasma levels of six quantified oxylipins decreased with the number of diseased arteries; a panel of five oxylipins diagnosed three diseased arteries with 100% sensitivity and 70% specificity. Concentrations of five oxylipins were lower and one oxylipin was higher with survival; a panel of two oxylipins predicted survival during follow-up with 86% sensitivity and 91% specificity.Conclusions: Quantification of plasma oxylipins may assist in CAD diagnosis and prognosis in combination with standard risk assessment tools.