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Over the years, there has been notable progress in understanding the pathogenesis and treatment modalities of diabetes and its complications, including the application of metabolomics in the study of diabetes, capturing attention from researchers worldwide. Advanced mass spectrometry, including gas chromatography–tandem mass spectrometry (GC-MS/MS), liquid chromatography–tandem mass spectrometry (LC-MS/MS), and ultra-performance liquid chromatography coupled to electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-Q-TOF-MS), etc., has significantly broadened the spectrum of detectable metabolites, even at lower concentrations. Advanced mass spectrometry has emerged as a powerful tool in diabetes research, particularly in the context of metabolomics. By leveraging the precision and sensitivity of advanced mass spectrometry techniques, researchers have unlocked a wealth of information within the metabolome. This technology has enabled the identification and quantification of potential biomarkers associated with diabetes and its complications, providing new ideas and methods for clinical diagnostics and metabolic studies. Moreover, it offers a less invasive, or even non-invasive, means of tracking disease progression, evaluating treatment efficacy, and understanding the underlying metabolic alterations in diabetes. This paper summarizes advanced mass spectrometry for the application of metabolomics in diabetes mellitus, gestational diabetes mellitus, diabetic peripheral neuropathy, diabetic retinopathy, diabetic nephropathy, diabetic encephalopathy, diabetic cardiomyopathy, and diabetic foot ulcers and organizes some of the potential biomarkers of the different complications with the aim of providing ideas and methods for subsequent in-depth metabolic research and searching for new ways of treating the disease. [ABSTRACT FROM AUTHOR]

As the ability to collect profiling data in metabolomics increases substantially with the advances in Liquid Chromatography–Mass Spectrometry (LC-MS) instruments, it is urgent to develop new and powerful data analysis approaches to match the big data collected and to extract as much meaningful information as possible from tens of thousands of molecular features. Here, we applied weighted gene co-expression network analysis (WGCNA), an algorithm popularly used in microarray or RNA sequencing, to plasma metabolomic data and demonstrated several advantages of WGCNA over conventional statistical approaches such as principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA). By using WGCNA, a large number of molecular features were clustered into a few modules to reduce the dimension of a dataset, the impact of phenotypic traits such as diet type and genotype on the plasma metabolome was evaluated quantitatively, and hub metabolites were found based on the network graph. Our work revealed that WGCNA is a very powerful tool to decipher, interpret, and visualize metabolomic datasets. [ABSTRACT FROM AUTHOR]

Brain tumors account for 1% of all cancers diagnosed de novo. Due to the specificity of the anatomical area in which they grow, they can cause significant neurological disorders and lead to poor functional status and disability. Regardless of the results of biochemical markers of intracranial neoplasms, they are currently of no diagnostic significance. The aim of the study was to use LC-ESI-MS/MS in conjunction with multivariate statistical analyses to examine changes in amino acid metabolic profiles between patients with glioblastoma, meningioma, and a group of patients treated for osteoarthritis of the spine as a control group. Comparative analysis of amino acids between patients with glioblastoma, meningioma, and the control group allowed for the identification of statistically significant differences in the amino acid profile, including both exogenous and endogenous amino acids. The amino acids that showed statistically significant differences (lysine, histidine, α-aminoadipic acid, phenylalanine) were evaluated for diagnostic usefulness based on the ROC curve. The best results were obtained for phenylalanine. Classification trees were used to build a model allowing for the correct classification of patients into the study group (patients with glioblastoma multiforme) and the control group, in which cysteine turned out to be the most important amino acid in the decision-making algorithm. Our results indicate amino acids that may prove valuable, used alone or in combination, toward improving the diagnosis of patients with glioma and meningioma. To better assess the potential utility of these markers, their performance requires further validation in a larger cohort of samples. [ABSTRACT FROM AUTHOR]

Ginsenoside Rg3, Rk1, and Rg5, rare ginsenosides from Panax ginseng, have many pharmacological effects, which have attracted extensive attention. They can be obtained through the heat treatment of Gynostemma pentaphyllum. In this study, scanning electron microscopy (SEM) and thermal gravity-differential thermal gravity (TG-DTG) were employed to investigate this process and the content change in ginsenosides was analyzed using liquid chromatography-mass spectrometry (LC-MS). SEM and TG-DTG were used to compare the changes in the ginsenosides before and after treatment. In SEM, the presence of hydrogen bond rearrangement was indicated by the observed deformation of vascular bundles and ducts. The before-and-after changes in the peak patterns and peaks values in TG-DTG indicated that the content of different kinds of compounds produced changes, which all revealed that the formation of new saponins before and after the heat treatment was due to the breakage or rearrangement of chemical bonds. Additionally, the deformation of vascular bundles and vessels indicated the presence of hydrogen bond rearrangement. The glycosidic bond at the 20 positions could be cleaved by ginsenoside Rb3 to form ginsenoside Rd, which, in turn, gave rise to ginsenoside Rg3(S) and Rg3(R). They were further dehydrated to form ginsenoside Rk1 and Rg5. This transformation process occurs in a weak acidic environment provided by G. pentaphyllum itself, without the involvement of endogenous enzymes. In addition, the LC-MS analysis results showed that the content of ginsenoside Rb3 decreased from 2.25 mg/g to 1.80 mg/g, while the contents of ginsenoside Rk1 and Rg5 increased from 0.08 and 0.01 mg/g to 3.36 and 3.35 mg/g, respectively. Ginsenoside Rg3(S) and Rg3(R) were almost not detected in G. pentaphyllum, and the contents of them increased to 0.035 and 0.23 mg/g after heat treatment. Therefore, the rare ginsenosides Rg3(S), Rg3(R), Rk1, and Rg5 can be obtained from G. pentaphyllum via heat treatment. [ABSTRACT FROM AUTHOR]

Kelussia odoratissima Mozaff. is a species of Apiaceae endemic to the Zagros Mountains in Iran. In the present investigation, for the first time, the polyphenolic compounds and flavonoids of its leaves were determined by liquid chromatography-mass spectrometry (LC-MS). As a result, p-coumaric acid, ferulic acid, caffeic acid, chlorogenic acid, acetyl phloroglucinol, vanillic acid, m-coumaric acid, and 4-methylsiringol were determined as the main phenolic compounds, while 3-hydroxyflavone, flavone, quercetin, rutin, neohesperidin, polydatin, and diosmin were the main flavonoid components, of which chlorogenic acid (303.08 µL/gDW), neohesperidin (38.37 µL/gDw), and diosmin (28.62 µL/gDW) were the most abundant. Solid-phase microextraction (SPME) was also used to determine the chemical compounds. Based on SPME, (Z)-undec-6-en-2-one (17.48%) and (Z)-butylidenephthalide (4.348%) were the major components. Based on GC-MS analyses, (Z)-ligustilide was the main compound; however, some new compounds were also determined, including 3-ethylisobenzofuran-1 (3H)-one, (E)-ligugustilide, and E-n-butylidene phthalide. Also, for the first time, we have identified EOs ethyl and isobutyl phthalides on the basis of the obtained EI-MS spectra. Finally, the fragmentation of phthalides is also discussed in this research. [ABSTRACT FROM AUTHOR]

The large quantity of olive roots resulting from a large number of old and unfruitful trees encouraged us to look for ways of adding value to these roots. For this reason, the current research work is devoted to the valorization of olive roots by identifying active phytochemicals and assessing their biological activities, including the cytotoxicity and antiviral potential of different extracts from the Olea europaea Chemlali cultivar. The extract, obtained by ultrasonic extraction, was analyzed using the liquid chromatography-mass spectrometry technique (LC-MS). The cytotoxicity was evaluated through the use of the microculture tetrazolium assay (MTT) against VERO cells. Subsequently, the antiviral activity was determined for HHV-1 (Human Herpesvirus type 1) and CVB3 (Coxsackievirus B3) replication in the infected VERO cells. LC-MS analysis allowed the identification of 40 compounds, classified as secoiridoids (53%), organic acids (13%), iridoids (10%), lignans (8%), caffeoylphenylethanoid (5%), phenylethanoids (5%),sugars and derivatives (2%), phenolic acids (2%), and flavonoids (2%). It was found that extracts were not toxic to the VERO cells. Moreover, the extracts did not influence the appearance of HHV-1 or CVB3 cytopathic effects in the infected VERO cells and failed to decrease the viral infectious titer. [ABSTRACT FROM AUTHOR]

In the quest to market increasingly safer and more potent biotherapeutic proteins, the concept of the multi-attribute method (MAM) has emerged from biopharmaceutical companies to boost the quality-by-design process development. MAM strategies rely on state-of-the-art analytical workflows based on liquid chromatography coupled to mass spectrometry (LC–MS) to identify and quantify a selected series of critical quality attributes (CQA) in a single assay. Here, we aimed at evaluating the repeatability and robustness of a benchtop LC–MS platform along with bioinformatics data treatment pipelines for peptide mapping-based MAM studies using standardized LC–MS methods, with the objective to benchmark MAM methods across laboratories, taking nivolumab as a case study. Our results evidence strong interlaboratory consistency across LC–MS platforms for all CQAs (i.e., deamidation, oxidation, lysine clipping and glycosylation). In addition, our work uniquely highlights the crucial role of bioinformatics postprocessing in MAM studies, especially for low-abundant species quantification. Altogether, we believe that MAM has fostered the development of routine, robust, easy-to-use LC–MS platforms for high-throughput determination of major CQAs in a regulated environment. [ABSTRACT FROM AUTHOR]

In the present study, lysozyme was purified by the following multi-step methodology: salt (ammonium sulfate) precipitation, dialysis, and ultrafiltration. The lysozyme potential was measured by enzymatic activity after each purification step. However, after ultrafiltration, the resulting material was considered extra purified. It was concentrated in an ultrafiltration centrifuge tube, and the resulting protein/lysozyme was used to determine its bactericidal potential against five bacterial strains, including three gram-positive (Bacillus subtilis 168, Micrococcus luteus, and Bacillus cereus) and two gram-negative (Salmonella typhimurium and Pseudomonas aeruginosa) strains. The results of ZOI and MIC/MBC showed that lysozyme had a higher antimicrobial activity against gram-positive than gram-negative bacterial strains. The results of the antibacterial activity of lysozyme were compared with those of ciprofloxacin (antibiotic). For this purpose, two indices were applied in the present study: antimicrobial index (AMI) and percent activity index (PAI). It was found that the purified lysozyme had a higher antibacterial activity against Bacillus cereus (AMI/PAI; 1.01/101) and Bacillus subtilis 168 (AMI/PAI; 1.03/103), compared to the antibiotic (ciprofloxacin) used in this study. Atomic force microscopy (AFM) was used to determine the bactericidal action of the lysozyme on the bacterial cell. The purified protein was further processed by gel column chromatography and the eluate was collected, its enzymatic activity was 21.93 U/mL, while the eluate was processed by native-PAGE. By this analysis, the un-denatured protein with enzymatic activity of 40.9 U/mL was obtained. This step shows that the protein (lysozyme) has an even higher enzymatic potential. To determine the specific peptides (in lysozyme) that may cause the bactericidal potential and cell lytic/enzymatic activity, the isolated protein (lysozyme) was further processed by the SDS-PAGE technique. SDS-PAGE analysis revealed different bands with sizes of 34 kDa, 24 kDa, and 10 kDa, respectively. To determine the chemical composition of the peptides, the bands (from SDS-PAGE) were cut, enzymatically digested, desalted, and analyzed by LC-MS (liquid chromatography-mass spectrometry). LC-MS analysis showed that the purified lysozyme had the following composition: the number of proteins in the sample was 56, the number of peptides was 124, and the number of PSMs (peptide spectrum matches) was 309. Among them, two peptides related to lysozyme and bactericidal activities were identified as: A0A1Q9G213 (N-acetylmuramoyl-L-alanine amidase) and A0A1Q9FRD3 (D-alanyl-D-alanine carboxypeptidase). The corresponding protein sequence and nucleic acid sequence were determined by comparison with the database. [ABSTRACT FROM AUTHOR]

Interest in cultivating cannabis for medical and recreational purposes is increasing due to a dramatic shift in cannabis legislation worldwide. Therefore, a comprehensive understanding of the composition of secondary metabolites, cannabinoids, and terpenes grown in different environmental conditions is of primary importance for the medical and recreational use of cannabis. We compared the terpene and cannabinoid profiles using gas/liquid chromatography and mass spectrometry for commercial cannabis from genetically identical plants grown indoors using artificial light and artificially grown media or outdoors grown in living soil and natural sunlight. By analyzing the cannabinoids, we found significant variations in the metabolomic profile of cannabis for the different environments. Overall, for both cultivars, there were significantly greater oxidized and degraded cannabinoids in the indoor-grown samples. Moreover, the outdoor-grown samples had significantly more unusual cannabinoids, such as C4- and C6-THCA. There were also significant differences in the terpene profiles between indoor- and outdoor-grown cannabis. The outdoor samples had a greater preponderance of sesquiterpenes including β-caryophyllene, α-humulene, α-bergamotene, α-guaiene, and germacrene B relative to the indoor samples. [ABSTRACT FROM AUTHOR]

A novel COVID-19 vaccine (BriLife®) has been developed by the Israel Institute for Biological Research (IIBR) to prevent the spread of the SARS-CoV-2 virus throughout the population in Israel. One of the components in the vaccine formulation is tris(hydroxymethyl)aminomethane (tromethamine, TRIS), a buffering agent. TRIS is a commonly used excipient in various approved parenteral medicinal products, including the mRNA COVID-19 vaccines produced by Pfizer/BioNtech and Moderna. TRIS is a hydrophilic basic compound that does not contain any chromophores/fluorophores and hence cannot be retained and detected by reverse-phase liquid chromatography (RPLC)-ultraviolet (UV)/fluorescence methods. Among the few extant methods for TRIS determination, all exhibit a lack of selectivity and/or sensitivity and require laborious sample treatment. In this study, LC–mass spectrometry (MS) with its inherent selectivity and sensitivity in the multiple reaction monitoring (MRM) mode was utilized, for the first time, as an alternative method for TRIS quantitation. Extensive validation of the developed method demonstrated suitable specificity, linearity, precision, accuracy and robustness over the investigated concentration range (1.2–4.8 mg/mL). Specifically, the R2 of the standard curve was >0.999, the recovery was >92%, and the coefficient of variance (%CV) was <12% and <6% for repeatability and intermediate precision, respectively. Moreover, the method was validated in accordance with strict Good Manufacturing Practice (GMP) guidelines. The developed method provides valuable tools that pharmaceutical companies can use for TRIS quantitation in vaccines and other pharmaceutical products. [ABSTRACT FROM AUTHOR]

Propolis, popularly known as bee glue, is a resinous, sticky substance produced by different bee species across the globe. Studies on the biological properties of propolis from the Philippines are rare. Hence, the current study aims at the chemical characterization of propolis produced by the stingless bees Tetragonula biroi Friese from the Philippines and to investigate its antitrypanosomal and anticancer properties. The determination of the chemical composition and characterization of propolis samples was achieved using liquid chromatography–mass spectrometry (LC-MS), -high-performance liquid chromatography–evaporative light scattering detector (HPLC-ELSD), and nuclear magnetic resonance (NMR) spectroscopy. Three major triterpenes were isolated and identified using HRESI-MS and 1H/13C NMR techniques. The spectral studies confirmed the presence of compounds such as isomangiferolic acid, 27-hydoxymangiferonic acid, and 27-hydroxyisomangiferolic acid. All crude propolis samples, isolated fractions, and pure compounds demonstrated moderate antitrypanosomal and anticancer properties compared to control drugs. Amongst the tested compounds, 27-hydoxymangiferonic acid exhibited the highest antitrypanosomal activity at a concentration of 11.6 µg/mL. The highest anticancer effect was demonstrated by the Ph-2 fraction, followed by 27-hydroxyisomangiferolic acid, with IC50 values of 129.6 and 153.3 µg/mL. Thus, it can be concluded that the observed biological activity of Philippine propolis is due to the combinatorial effect or synergistic action of the active compounds 27-hydoxymangiferonic acid and 27-hydroxyisomangiferolic acid. [ABSTRACT FROM AUTHOR]

Antibody–drug conjugates (ADCs) are a new class of biotherapeutics, consisting of a cytotoxic payload covalently bound to an antibody by a linker. Ligand-binding assay (LBA) and liquid chromatography-mass spectrometry (LC-MS) are the favored techniques for the analysis of ADCs in biomatrices. The goal of our review is to provide current strategies related to a series of bioanalytical assays for pharmacokinetics (PK) and anti-drug antibody (ADA) assessments. Furthermore, the strengths and limitations of LBA and LC-MS platforms are compared. Finally, potential factors that affect the performance of the developed assays are also provided. It is hoped that the review can provide valuable insights to bioanalytical scientists on the use of an integrated analytical strategy involving LBA and LC–MS for the bioanalysis of ADCs and related immunogenicity evaluation. [ABSTRACT FROM AUTHOR]

This study was designed to evaluate the chemical fingerprints and the antioxidant, anti-inflammatory and hypolipidemic activity of selected Crepis species collected in Greece, namely, C. commutata, C. dioscoridis, C. foetida, C. heldreichiana, C. incana, C. rubra, and Phitosia crocifolia (formerly known as Crepis crocifolia). For the phytochemical analyses, sample measurements were carried out by using nuclear magnetic resonance (NMR) spectroscopy and liquid chromatography coupled with mass spectrometry (LC-MS). Τhe extracts were evaluated both in vitro (radical scavenging activity: DPPH assay and total phenolic content: Folin–Ciocalteu) and in vivo (paw edema reduction and hypolipidemic activity: experimental mouse protocols). Among the tested extracts, C. incana presented the highest gallic acid equivalents (GAE) (0.0834 mg/mL) and the highest antioxidant activity (IC50 = 0.07 mg/mL) in vitro, as well as the highest anti-inflammatory activity with 32% edema reduction in vivo. Moreover, in the hypolipidemic protocol, the same extract increased plasma total antioxidant capacity (TAC) by 48.7%, and decreased cholesterol (41.3%) as well as triglycerides (37.2%). According to fractionation of the extract and the phytochemical results, this biological effect may be associated with the rich phenolic composition; caffeoyl tartaric acid derivatives (cichoric and caftaric acid) are regarded as the most prominent bioactive specialized metabolites. The present study contributes to the knowledge regarding the phytochemical and pharmacological profile of Crepis spp. [ABSTRACT FROM AUTHOR]

Background: Commercial pharmaceutical herbal products have enabled people to take traditional Chinese medicine (TCM) in a convenient and accessible form. However, the quantity and quality should be additionally inspected. To address the issue, a combination of chemical and physical inspection methods were developed to evaluate the amount of an herbal formula, Xiang-Sha-Liu-Jun-Zi-Tang (XSLJZT), in clinical TCM practice. Methods: A high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS) method with electrospray ionization was developed to measure the herbal biomarkers of guanosine, atractylenolide III, glycyrrhizic acid, dehydrocostus lactone, hesperidin, and oleanolic acid from XSLJZT. Scanning electron microscopy (SEM) photographs and light microscopy photographs with Congo red and iodine-KI staining were used to identify the cellulose fibers and starch content. Furthermore, solubility analysis, swelling power test, and crude fiber analysis were contributed to measure the starch additive in pharmaceutical products. Results: The results demonstrated large variations in the chemical components of different pharmaceutical brands. The SEM photographs revealed that the starch was oval, smooth, and granular, and that the raw herbal powder appears stripy, stretched, and filiform. The stained light microscopy photographs of all of the pharmaceutical products showed added starch and raw herbal powder as extenders. Conclusion: The developed chemical and physical methods provide a standard operating procedure for the quantity control of the herbal pharmaceutical products of XSLJZT. [ABSTRACT FROM AUTHOR]

Catharanthus roseus (C. roseus) and Vinca minor (V. minor) are two common important medical plants belonging to the family Apocynaceae. In this study, we used non-targeted GC-MS and targeted LC-MS metabolomics to dissect the metabolic profile of two plants with comparable phenotypic and metabolic differences. A total of 58 significantly different metabolites were present in different quantities according to PCA and PLS-DA score plots of the GC-MS analysis. The 58 identified compounds comprised 16 sugars, eight amino acids, nine alcohols and 18 organic acids. We subjected these metabolites into KEGG pathway enrichment analysis and highlighted 27 metabolic pathways, concentrated on the TCA cycle, glycometabolism, oligosaccharides, and polyol and lipid transporter (RFOS). Among the primary metabolites, trehalose, raffinose, digalacturonic acid and gallic acid were revealed to be the most significant marker compounds between the two plants, presumably contributing to species-specific phenotypic and metabolic discrepancy. The profiling of nine typical alkaloids in both plants using LC-MS method highlighted higher levels of crucial terpenoid indole alkaloid (TIA) intermediates of loganin, serpentine, and tabersonine in V. minor than in C. roseus. The possible underlying process of the metabolic flux from primary metabolism pathways to TIA synthesis was discussed and proposed. Generally speaking, this work provides a full-scale comparison of primary and secondary metabolites between two medical plants and a metabolic explanation of their TIA accumulation and phenotype differences. [ABSTRACT FROM AUTHOR]

Salvia miltiorrhiza (S. miltiorrhiza) Bunge is broadly used as herbal medicine for the clinical treatments of cardiovascular and cerebrovascular diseases. Despite its commercial and medicinal values, few systematic studies on the metabolome of S. miltiorrhiza roots have been carried out so far. We systematically described the metabolic profiles of S. miltiorrhiza using high pressure liquid chromatography mass spectrometry (LC/MS) in conjunction with multivariate statistical analyses, aimed at monitoring their biological variations of secondary metabolites related to three locations and four S. miltiorrhiza genotypes. A total of 40 bioactive constituents were putatively annotated in S. miltiorrhiza root samples. This study found that both the same S. miltiorrhiza genotype growing at three different locations and four S. miltiorrhiza genotypes growing at the same location had significant metabonomic differences identified by the principal component analysis (PCA) approach. By using orthogonal projection to latent structure with discriminant analysis (OPLS-DA), 16 and 14 secondary metabolites can be used as potential location-specific and genotype-specific markers in S. miltiorrhiza, respectively. The specificity of LC/MS profiles offered a powerful tool to discriminate S. miltiorrhiza samples according to genotypes or locations. [ABSTRACT FROM AUTHOR]

The present work was performed to investigate the phenolic composition of P. lentiscus L. distilled leaves (PDL) and examine its potential against certain key enzymes related to skin aging. High-pressure liquid chromatography coupled to mass spectrometry (HPLC-MS) and various separation procedures combined with nuclear magnetic resonance (NMR) and MS analysis were performed to isolate and identify compounds present in the ethyl acetate extract (EAE) of PDL. A high amount of flavonol glycoside was detected in EAE. Indeed, quercetin-3-O-rhamnoside (FC), myricetin-3-O-rhamnoside (FM2), and kaempferol-3-O-rhamnoside (FB2) were isolated from EAE, and are present in high quantities of 10.47 ± 0.26, 12.17 ± 0.74, and 4.53 ± 0.59 mg/g dry weight, respectively. A transdermal diffusion study was carried out to determine the EAE-molecules that may transmit the cutaneous barrier and showed that FM2 transmits the membrane barrier with a high amount followed by FC. EAE, FM2, and FC were tested against tyrosinase and elastase enzymes. Moreover, intracellular tyrosinase inhibition and cytotoxicity on skin melanoma cells (B16) were evaluated. The results indicated that EAE, FC, and FM2 have important inhibitory activities compared to the well-known standards, at non-cytotoxic concentrations. Therefore, they could be excellent agents for treating skin pigmentation and elasticity problems. [ABSTRACT FROM AUTHOR]

The main objective of the current work was to evaluate the suitability of ultrasound-assisted extraction (UAE) for the recovery of oil from papaya seed as compared to conventional extraction techniques (i.e., Soxhlet extraction (SXE) and solvent extraction (SE)). In the present study, the recovery yield, fatty acid composition and triacylglycerol profile of papaya seed oil obtained from different extraction methods and conditions were compared. Results indicated that both solvent extraction (SE, 12 h/25 °C) and ultrasound-assisted extraction (UAE) methods recovered relatively high yields (79.1% and 76.1% of total oil content, respectively). Analysis of fatty acid composition revealed that the predominant fatty acids in papaya seed oil were oleic (18:1, 70.5%-74.7%), palmitic (16:0, 14.9%-17.9%), stearic (18:0, 4.50%-5.25%), and linoleic acid (18:2, 3.63%-4.6%). Moreover, the most abundant triacylglycerols of papaya seed oil were triolein (OOO), palmitoyl diolein (POO) and stearoyl oleoyl linolein (SOL). In this study, ultrasound-assisted extraction (UAE) significantly (p < 0.05) influenced the triacylglycerol profile of papaya seed oil, but no significant differences were observed in the fatty acid composition of papaya seed oil extracted by different extraction methods (SXE, SE and UAE) and conditions. [ABSTRACT FROM AUTHOR]

Current methods for measuring the abundance of proteogenic amino acids in plants require derivatisation, extended run times, very sensitive pH adjustments of the protein hydrolysates, and the use of buffers in the chromatographic phases. Here, we describe a fast liquid chromatography–mass spectrometry (LC–MS) method for the determination of amino acids that requires only three steps: hydrolysis, neutralisation, and sample dilution with a borate buffer solution for pH and retention time stability. The method shows excellent repeatability (repeated consecutive injections) and reproducibility (repeated hydrolysis) in the amino acid content, peak area, and retention time for all the standard amino acids. The chromatographic run time is 20 min with a reproducibility and repeatability of <1% for the retention time and <11% for the peak area of the BSA and quality control (QC) lentil samples. The reproducibility of the total protein levels in the hydrolysis batches 1–4 was <12% for the BSA and the lentil samples. The level of detection on column was below 0.1 µM for most amino acids (mean 0.017 µM). [ABSTRACT FROM AUTHOR]

Early detection of any preeclampsia biomarkers may lower the risk of mortality, both for a mother and a child. Our study focuses on techniques for preeclampsia biomarker identification by comparing the results of a method using liquid chromatography mass spectrometry in multiple reaction monitoring mode (LC-MS/MS) with those by the enzyme-linked immunosorbent assay (ELISA) test, as well as by comparing the obtained results with clinical data. In the proposed LC-MS/MS method a tryptic digest peptide charge derivatization strategy was used as a tool for sensitive detection of podocin, i.e., a previously discovered preeclampsia biomarker present in urine samples from pregnant women. Urine samples from pregnant women with diagnosed preeclampsia were collected at different stages of pregnancy and from healthy subjects, and then were analyzed by ELISA test and the proposed method with LC-MS/MS. Charge derivatization of the ε amino group of C-terminal lysine residues in tryptic digests by 2,4,6-triphenylpyrylium salt was performed to increase the ionization efficiency in the LC-MS/MS mode. Podocin was identified at the early stage of pregnancy, while its detection using an ELISA test was not possible. The protocol for urine sample preparation was optimized. Our results show that the proposed method by LC-MS/MS in combination with peptide charge derivatization, provides an ultrasensitive tool for diagnosis of preeclampsia, and provides earlier detection than a clinical diagnosis or ELISA test. The proposed solution may revolutionize medical diagnostics. [ABSTRACT FROM AUTHOR]

Bopu powder® and Sangrovit® were developed from Macleayacordata and are widely used in agriculture and animal husbandry, but their impurities have been rarely reported in the literature. Impurity analysis is of great importance to the quality and safety of veterinary drugs. In this study, high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS) combined with a screening method was used to screen and characterize the impurities in Bopu powder® and Sangrovit®. A total of 58 impurities were screened from Bopu powder® and Sangrovit® using the screening strategies, of which 39 were identified by their accurate m/z value, characteristic MS/MS data, and fragmentation pathways of references. This established method was used for impurity analysis for the first time and proved to be a useful and rapid tool to screen and identify the impurities of Bopu powder® and Sangrovit®, especially for those at trace levels in a complex sample. In addition, this study marks the first comprehensive research into impurities in these two products and has great significance for the systematic detection of impurities in other plant-derived drugs. [ABSTRACT FROM AUTHOR]

Fermented aqueous extracts of Viscum album L. are widely used for cancer treatment in complementary medicine. The high molecular weight compounds viscotoxins and lectins are considered to be the main active substances in the extracts. However, a vast number of small molecules (≤1500 Da) is also expected to be present, and few studies have investigated their identities. In this study, a comprehensive metabolome analysis of samples of fermented aqueous extracts of V. album from two host tree species (Malus domestica and Pinus sylvestris), both prepared by two pharmaceutical manufacturing processes, was performed by liquid chromatography−high resolution tandem mass spectrometry (LC-HRMS/MS). A total of 212 metabolites were putatively annotated, including primary metabolites (e.g., amino acids, organic acids, etc.) and secondary metabolites (mostly phenolic compounds). A clear separation between V. album samples according to the host tree species, but not due to manufacturing processes, was observed by principal component analysis. The biomarkers responsible for this discrimination were assessed by partial least squares−discriminant analysis. Because V. album extracts from different host trees have different clinical applications, the present work highlights the possibility of characterizing the metabolome for identification and traceability of V. album fermented aqueous extracts. [ABSTRACT FROM AUTHOR]

Dicamba is a moderately volatile herbicide used for post-emergent control of broadleaf weeds in corn, soybean, and a number of other crops. With increased use of dicamba due to the release of dicamba-resistant cotton and soybean varieties, growing controversy over the effects of spray drift and volatilization on non-target crops has increased the need for quantifying dicamba collected from water and air sampling. Therefore, this study was designed to evaluate stable isotope-based direct quantification of dicamba from air and water samples using single-quadrupole liquid chromatography–mass spectrometry (LC–MS). The sample preparation protocols developed in this study utilize a simple solid-phase extraction (SPE) protocol for water samples and a single-step concentration protocol for air samples. The LC–MS detection method achieves sensitive detection of dicamba based on selected ion monitoring (SIM) of precursor and fragment ions and relies on the use of an isotopically labeled internal standard (IS) (D3-dicamba), which allows for calculating recoveries and quantification using a relative response factor (RRF). Analyte recoveries of 106–128% from water and 88–124% from air were attained, with limits of detection (LODs) of 0.1 ng mL−1 and 1 ng mL−1, respectively. The LC–MS detection method does not require sample pretreatment such as ion-pairing or derivatization to achieve sensitivity. Moreover, this study reveals matrix effects associated with sorbent resin used in air sample collection and demonstrates how the use of an isotopically labeled IS with RRF-based analysis can account for ion suppression. The LC–MS method is easily transferrable and offers a robust alternative to methods relying on more expensive tandem LC–MS/MS-based options. [ABSTRACT FROM AUTHOR]

The aim of this research was to examine the effect of the hydroalcoholic extracts from the peel (APE) and pulp (APP) of a traditional apple cultivar from central Italy (Mela Rosa dei Monti Sibillini) on CCl4-induced hepatotoxicity in rats. Phytoconstituents were determined by liquid chromatography–mass spectrometry (LC-MS) analysis showing an abundance of proanthocyanidins and flavonol derivatives together with the presence of annurcoic acid in APE. Wistar rats received APE/APP (30 mg/kg oral administration) for three days before CCl4 injection (2 mL/kg intraperitoneal once on the third day). Treatment with both APE and APP prior to CCl4 injection significantly decreased the serum levels of aspartate aminotransferase (AST), alkaline phosphatase (ALP) and alanine aminotransferase (ALT) compared to the CCl4 group. Besides, pretreatment with APE reversed the CCl4 effects on superoxide dismutase (SOD), myeloperoxidase (MPO), tumor necrosis factor-α (TNF-α) and interleukin-1beta (IL-1β) levels in liver tissue in rats and reduced tissue damage as shown in hematoxylin and eosin staining. These results showed that this ancient Italian apple is worthy of use in nutraceuticals and dietary supplements to prevent and/or protect against liver disorders. [ABSTRACT FROM AUTHOR]

Conventional fluorescent lamps that are used in tissue culture are costly light sources, exhibiting excessive wavelength emission-bandwidth that must be replaced by alternative, less costly, and much lower power-consuming energy sources. The use of Light-Emitting Diodes (LEDs) is the best option due to their potential role as elicitors of secondary metabolite production in many plant models. Gynura procumbens (G. procumbens) is widely used for treating various diseases. Here, leaf explants were cultivated in MS medium that was supplemented with 0.5 mg/L of naphthaleneacetic acid (NAA) and 2.0 mg/L of benzylaminopurine (BAP) for 30 days under white, blue, and red LEDs. Secondary metabolites were analyzed by High Performance Liquid Chromatography (HPLC) and Liquid Chromatography-Mass Spectrometry (LC-MS). Blue LEDs elicited the highest antioxidant activity, total flavonoid, and phenolic content. Furthermore, the content of cyanidin-monoglucosides significantly increased under blue light. [ABSTRACT FROM AUTHOR]

Nitrated lipids have been detected in vitro and in vivo, usually associated with a protective effect. While nitrated fatty acids have been widely studied, few studies reported the nitration and nitroxidation of the phospholipid classes phosphatidylcholine, and phosphatidylethanolamine. However, no information regarding nitrated and nitroxidized phosphatidylserine can be found in the literature. This work aims to identify and characterize the nitrated and nitroxidized derivatives of 1-palmitoyl-2-oleoyl-sn-3-glycero-phosphoserine (POPS), obtained after incubation with nitronium tetrafluoroborate, by liquid chromatography (LC) coupled to mass spectrometry (MS) and tandem MS (MS/MS). Several nitrated and nitroxidized products were identified, namely, nitro, nitroso, nitronitroso, and dinitro derivatives, as well as some nitroxidized species such as nitrosohydroxy, nitrohydroxy, and nitrohydroperoxy. The fragmentation pathways identified were structure-dependent and included the loss of HNO and HNO2 for nitroso and nitro derivatives, respectively. Combined losses of PS polar head group plus HNO or HNO2 and carboxylate anions of modified fatty acyl chain were also observed. The nitrated POPS also showed antiradical potential, demonstrated by the ability to scavenge the ABTS●+ and DPPH● radicals. Overall, this in vitro model of nitration based on LC-MS/MS provided additional insights into the nitrated and nitroxidized derivatives of PS and their fragmentation fingerprinting. This information is a valuable tool for targeted analysis of these modified PS in complex biological samples, to further explore the new clues on the antioxidant potential of nitrated POPS. [ABSTRACT FROM AUTHOR]

Tomato crops suffer attacks of various pathogens that cause large production losses. Late blight caused by Phytophthora infestans is a devastating disease in tomatoes because of its difficultly to control. Here, we applied metabolomics based on liquid chromatography–mass spectrometry (LC-MS) and metabolic profiling by matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) in combination with multivariate data analysis in the early detection of late blight on asymptomatic tomato plants and to discriminate infection times of 4, 12, 24, 36, 48, 60, 72 and 96 h after inoculation (hpi). MALDI-MS and LC-MS profiles of metabolites combined with multivariate data analysis are able to detect early-late blight-infected tomato plants, and metabolomics based on LC-MS discriminates infection times in asymptomatic plants. We found the metabolite tomatidine as an important biomarker of infection, saponins as early infection metabolite markers and isocoumarin as early and late asymptomatic infection marker along the post infection time. MALDI-MS and LC-MS analysis can therefore be used as a rapid and effective method for the early detection of late blight-infected tomato plants, offering a suitable tool to guide the correct management and application of sanitary defense approaches. LC-MS analysis also appears to be a suitable tool for identifying major metabolites of asymptomatic late blight-infected tomato plants. [ABSTRACT FROM AUTHOR]