Search Results (4,367)

In this study, an ion-pair reverse-phase high-performance liquid chromatography–electrospray ionization mass spectrometry (RP-HPLC-ESI-MS) method was optimized and validated for purity determination for the quality control of the proposed generic nusinersen oligonucleotide drug substance and drug product. The optimization and considerations of sample preparation, chromatographic and mass spectrometry conditions are discussed. The limit of detection was 2.5 × 10⁻⁵ mg/mL and the limit of quantitation was 4.9 × 10⁻⁵ mg/mL. The linearity of the signal (XIC) for all impurities was linear with correlation coefficients of R² ≥ 0.9669. This study, associated with the development of therapeutic oligonucleotides, examines the subject of product-related impurities. The authors consider an ion-pair reverse-phase high-performance liquid chromatography in combination with mass spectrometry for impurity quantitative control. This study contributes to the field by elucidating several critical aspects that, while previously unaddressed in the existing literature, are essential for developing effective analytical methods. Full article

Human infertility affects approximately 17.5% of the global population, with male factors accounting for nearly half of all cases. Identifying reliable molecular biomarkers is crucial for improving the diagnosis and assessment of male fertility. This study established and refined an untargeted high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC-ESI-MS/MS) protocol for a comprehensive lipidomic and metabolomic analysis of human spermatozoa, using only 1.25 million cells per sample. Compared with previous reports, our optimized method achieved an unparalleled level of analytical depth, identifying 473 lipid species and 955 structurally annotated metabolites. This corresponds to nearly a 7600-fold improvement in detection efficiency per cell compared with previously published approaches. Lipidomic analysis revealed that the most abundant lipid classes were glycerophospholipids (39%), cholesterol (20%) and fatty acids (19%), with cholesterol representing the single most abundant compound. This observation is consistent with the structural complexity of the sperm plasma membrane. Metabolomic profiling similarly identified glycerophospholipids (44%), eicosanoids (14%) and N-acyl amino acids (12%) as the major metabolite classes. The integration of lipidomic and metabolomic data highlighted functionally interconnected pathways related to membrane dynamics, energy metabolism, and hormone biosynthesis. Overall, this work establishes a robust, sensitive, and scalable analytical framework that enables the high-coverage molecular characterization of spermatozoa from limited sample material, laying the groundwork for future biomarker discovery and clinical applications in male infertility research. Full article

Glycidol, a probable human carcinogen, remains an under-investigated process contaminant in soy sauce. This study developed a sensitive and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for its determination in this complex condiment. The approach combined chemical derivatization with p-Dimethylaminophenol hydrochloride for analyte stabilization with an optimized sample pretreatment using a custom-packed activated carbon solid-phase extraction (SPE) cartridge effectively removed matrix interferences, and performing the derivatization at pH 6.5 prevented conversion of 2- and 3-monochloropropanediol (2-MCPD and 3-MCPD) into glycidol, ensuring high specificity and accuracy. This approach shows broad linearity from 1 to at least 100 ng/mL (R² = 0.9993), and demonstrates excellent performance, with a limit of detection and quantification of 0.5 ng/mL and 1.0 ng/mL, respectively. Application to commercial samples (n = 11) confirmed the presence of glycidol, highlighting the need for its monitoring. This work provides a robust analytical tool essential for supporting food safety surveillance of this contaminant in fermented foods. Full article

Alnus glutinosa (L.) Gaertn. has extensive use in traditional medicine and diverse biological activities due to its rich phytochemical profile. In this study, firstly, the physicochemical characteristics of the plant material were evaluated, revealing a high content of extractive substances (17.684%), followed by ash (6.740%) and moisture (5.000%). Among the bioactive constituents, tannins were the most abundant (7.439%). Analysis of macroelements in the plant ash showed K (11.4330 mg/g) as the predominant element, followed by Mg (97.13 mg/g), Ca (75.30 mg/g), and Na (72.41 mg/g). Trace element analysis indicated Fe (1.2266 mg/g) as the most abundant microelement, with Zn (0.8870 mg/g) and Mn (0.8141 mg/g) present in comparable amounts. Gas chromatography–mass spectrometry (GC-MS) analysis of the ethanolic leaf extract characterized volatile and semi-volatile constituents of 43 phytochemical components, where vitamin E was the predominant compound (20.52%), followed by phytol (12.46%) and squalene (10.29%). Further high-performance liquid chromatography (HPLC) analysis confirmed the presumed presence of naringin (56.421 mg/L), followed by epicatechin (15.123 mg/L), catechin (12.485 mg/L), and phloridzin (11.800 mg/L), while gallic acid was detected at a comparatively lower concentration (0.402 mg/L). The antimicrobial activity of the aqueous leaf extract was evaluated against typical Gram-positive and Gram-negative pathogens, including Staphylococcus aureus, Salmonella abony, Escherichia coli, and Klebsiella pneumoniae. To evaluate the effect of compositional changes on antimicrobial activity, the fermented and non-fermented formulations of A. glutinosa leaf extracts were prepared. These results demonstrate measurable antibacterial effects, thereby confirming the ethnopharmacological significance of A. glutinosa and highlighting its potential as a source of natural antimicrobial agents for further pharmacological development. Full article

Phenolic compounds are plant-derived antioxidants crucial for human health and food preservation. Their bioactive potential including anti-inflammatory, antimicrobial, and anti-carcinogenic properties makes them a vital focus in nutritional, pharmaceutical, and agricultural research. This review critically evaluates the methodologies for their extraction, detection, and quantification to accurately assess antioxidant activity. Oxidative stress in biological systems and food matrices necessitates accurate analytical methodologies for assessing antioxidant behavior, which include both in vitro, in vivo and ex vivo approaches. Sample pretreatment and extraction techniques are critical for reliable analysis and vary depending on the matrix, compound polarity, and target phenolic subclass. We compare conventional extraction techniques (Soxhlet, maceration) with advanced methods like ultrasound-assisted, microwave-assisted, and supercritical fluid extraction. Detection methods reviewed include spectrophotometric assays (e.g., DPPH, FRAP, ORAC), electrochemical sensors, and chromatographic techniques (e.g., HPLC, HPLC−MS). While each method has distinct advantages, a lack of standardization remains the primary challenge, driven by variations in protocols and the vast chemical diversity of phenolics. This review underscores the critical need for integrated, standardized approaches to ensure the accurate and comparable evaluation of antioxidant activity in research and industry. Full article

Extra virgin olive oil (EVOO) is a key component of the Mediterranean diet, valued for its bioactive constituents and associated health benefits. This study evaluated the influence of four agronomic and processing factors—harvest month, destoning, fruit washing, and bottling delay—on the chemical composition of Kolovi EVOOs from the PGI Lesvos region. A total of 34 oils were produced under standardized conditions and analyzed for phenolic compounds, tocopherols, pigments, and squalene using UPLC-QTOF-MS and HPLC-DAD. The oils were characterized by consistently high nutritional quality, with most samples fulfilling EFSA health claim thresholds for hydroxytyrosol, tyrosol and its derivatives, and α-tocopherol. Harvest month was the most influential parameter: early harvested oils (October) contained significantly higher levels of phenolics, α-tocopherol, and lutein, whereas later harvests (November) were richer in squalene. Destoning produced modest changes, with slightly higher phenolics in non-destoned oils and reduced lipophilic antioxidants in destoned samples. Fruit washing selectively decreased hydrophilic phenolics, while lipophilic compounds were largely unaffected. Bottling delays of up to 48 h under protective conditions had negligible effects on composition, aside from minor increases in specific phenolic derivatives. These findings suggest that early harvesting and careful consideration of destoning are the most effective strategies for supporting the antioxidant profile of Kolovi EVOOs, while other practices can be adjusted with limited impact on quality. Full article

In the present work, the blackening process of Panax quinquefolius L. (PQ) was systematically investigated at temperatures of 70–90 °C, relative humidities (RHs) of 70–85%, and treatment times of 0–14 days. Ginsenoside compositions and transformation pathways were analyzed by high-performance liquid chromatography (HPLC) and liquid chromatography coupled with ion trap time-of-flight tandem mass spectrometry (LC-IT-TOF-MS/MS). The results demonstrated that blackening treatment significantly increased total saponin content from 2.72% to 5.73% after being treated at 80 °C and 70% RH for 12 days, accompanied by the highest conversion efficiencies for newly generated ginsenosides Rk1 (8.89 mg/g) and Rg5 (17.69 mg/g). Furthermore, compared with untreated PQ saponins (PQS), the blackened PQ saponins treated under optimal conditions (BPQS) exhibited superior 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) cation (ABTS⁺) radical scavenging activities, with IC₅₀ values of 0.2999 mg/mL and 0.2640 mg/mL, respectively, as well as stronger reducing power. Meanwhile, BPQS exhibited higher cytotoxicity toward HepG2 cells and effectively inhibited cell survival and proliferation by promoting the expression of apoptosis-related proteins, including caspase 3 and caspase 9. Our findings indicate that BPQS may be a functional ingredient suitable for use in dietary supplements and disease chemoprevention. Full article

Aim: This study investigates how Buyang Huanwu Decoction (BHD) protects against cerebral ischemic damage by targeting the NLRP3/Caspase-1 pathway. Methods: The fingerprint of BHD was analyzed by HPLC-UV. Migratory chemicals in BHD-containing cerebrospinal fluid (BHD-CCSF) were analyzed by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS). The effects of BHD on the NLRP3/Caspase-1 pathway, IL-18 and IL-1β levels in oxygen and glucose deprivation/reperfusion (OGD/R) cells were assessed by Western blot and ELISA. Cerebral infarction severity in permanent middle cerebral artery occlusion (pMCAO) mice was assessed by mNSS scores and staining. Protein and mRNA levels of the NLRP3/Caspase-1 pathway and inflammatory factors (IL-18, IL-1β) were measured. Results: BHD-containing serum (BHD-CS), BHD-CCSF, and Calycosin (Cal) reduced NLRP3, Caspase-1, ASC, GSDMD proteins, IL-18 and IL-1β in OGD/R cells. In pMCAO mice, BHD decreased pathway-related proteins and mRNA and inflammatory factors and alleviated brain injury. Conclusions: BHD ameliorates cerebral ischemia by inhibiting the NLRP3/Caspase-1 pathway, thereby suppressing pyroptosis and inflammation. Full article

Randia echinocarpa is an endemic shrub species of Mexico, commonly known as papache in the state of Sinaloa, where it has traditionally been used in medicinal practices. The present study evaluated the phenolic profile and antioxidant capacity of different tissues (leaf, bark, and fruit pulp) of R. echinocarpa. Phenolic compounds were characterized using HPLC–PDA–MS, which allowed the identification of seven compounds in the leaf, six in the bark, and six in the fruit pulp. Chlorogenic acid, ellagic acid, and rutin were among the most abundant compounds detected. Total phenolic content varied depending on tissue and season, with the highest concentration observed in leaves during autumn (2.770 ± 0.011 mg GAE g⁻¹) and the lowest in bark during winter (0.437 ± 0.009 mg GAE g⁻¹). This study also reports, for the first time, the concentrations of tannins and flavonoids in R. echinocarpa, with the highest content found in leaves during autumn (0.261 ± 0.003 mg EE g⁻¹ and 2.186 ± 0.005 mg RE g⁻¹, respectively). Antioxidant capacity was evaluated using DPPH and ABTS radical scavenging assays, with leaf extracts showing the highest activity, with IC₅₀ values of 0.82 mg mL⁻¹ and 1.21 mg mL⁻¹, respectively. These results provide new information on the phenolic composition and antioxidant potential of R. echinocarpa, contributing to the phytochemical characterization of this traditionally used medicinal species. Full article

Genome sequencing has revealed that microorganisms have the potential to produce many more natural products than previously thought; the challenge is to establish efficient ways to “wake up” those “sleeping” biosynthetic pathways or genes, which are undoubtedly expressed in nature under specific conditions that are not normally reproduced in the laboratory. To activate these cryptic natural products, co-cultivation of cross-kingdom microorganisms between Candida albicans and Streptomyces longshengensis was performed in this study. A novel peak generated through co-culture was isolated and analyzed by a high-performance liquid chromatograph (HPLC), and its chemical structure was further determined by using mass spectrum (MS) and nuclear magnetic resonance (NMR) analyses. Bioassays of antimicrobial and antitumor activities were performed, and heterologous expression in Escherichia coli was attempted. The chemical structure was identified as tryptophol, and the bioassays revealed that tryptophol showed antitumor activity with IC₅₀ values of 154.5, 144.3, 122.6, and 110.7 μg/mL against A549, MC38, HepG2, and MCF-7 cells, respectively. As a valuable compound, tryptophol was also heterologously expressed in E. coli C41 to address the drawbacks of chemical synthesis. These findings combine co-cultivation with genetic engineering for tryptophol biosynthesis, expanding its antitumor application and laying a foundation for its industrial and sustainable production. Full article

This study investigated the influence of cocoa-bean amino acid profile on the formation of α,β-unsaturated carbonyls. Some of these heat-derived compounds generated primarily during roasting (often through Strecker degradation and aldol condensation) are of toxicological concern (e.g., the aneugenic 2-phenylcrotonaldehyde and the genotoxic furan-2(5H)-one). Here, their levels were compared by SAFE-GC/MS in cocoa of different origins, before and after roasting. Free amino acid profiles were determined by HPLC-MS/MS. All the investigated roasted beans showed similar total free amino acid contents (10.5–15.0 g·kg⁻¹) and profiles, while α,β-unsaturated aldehyde levels differed markedly between samples, and reached >200 µg·kg⁻¹ after roasting, indicating that amino acid availability alone does not govern their formation. Analysis of a commercial cocoa-free chocolate surrogate, however, revealed much lower amounts of both free amino acids (total 1.6 g·kg⁻¹) and amino-acid-derived α,β-unsaturated aldehydes (total 11 µg·kg⁻¹) than in chocolate, together with only traces of pyrazines (total 72 µg·kg⁻¹). In contrast, furan-2(5H)-one was found at similar levels in chocolate and the cocoa-free product (56–79 µg·kg⁻¹), confirming a completely different pathway where amino acids are not key reagents in its synthesis. Full article

The hematite phase decorated with iron-doped cerium oxide nanoparticles (F@FC) was precipitated from cerium and iron oxalate intermediate products. The photocatalytic composite of graphitic carbon nitride (gCN) and F@FC was prepared by a simple method involving mixing the two components, followed by thermal treatment at 400 °C. According to electron microscopy, F@FC is composed of a submicron iron oxide (hematite) phase decorated with iron-doped cerium oxide nanoparticles deposited on gCN substrate. A hierarchically structured composite was observed instead of a simple mechanical mixture of α-Fe₂O₃, Fe-CeO₂, and gCN. To observe two types of degradation activity, photocatalytic and Photo-Fenton degradation activity, Rhodamine B (RhB) was applied as the model water pollutant. The influence of the amount of photocatalyst, the RhB concentration, the presence of cations and anions, the pH, and the effect of e⁻, h⁺, •OH, and •O₂⁻ scavenging reactants were studied. The Photo-Fenton degradation exhibited high efficiency across the entire tested pH range, whereas photocatalytic degradation showed comparable activity only at acidic pH. The F@FC-gCN composite catalyst exhibited a high degree of recyclability. The degradation pathways of photocatalytic and Photo-Fenton reactions were suggested by HPLC-MS analysis of the reaction products. A notable finding of this study was the observation that the green-yellow, fluorescent intermediate Rhodamine 110 was formed during the photocatalytic degradation of RhB. However, the high reactivity of the generated •OH radicals during Photo-Fenton degradation has been demonstrated to inhibit the formation of intermediate Rhodamine 110. Full article

The chemical composition of Iris songarica rhizome extracts was systematically investigated using GC-MS and UHPLC-MS. Their biological activity was further evaluated in vivo. The chloroform rhizome extract contained 33 identified compounds distributed across five main classes. Flavonoids predominated (50.7% of total ionic current), with tectochrysine (42.15%) as the major component, followed by 3,7-dihydroxy-2-(3,4-dimethoxyphenyl)-4H-chromene-4-one (5.18%) and a naringenin derivative (3.99%). Fatty acid esters comprised 30.6%, dominated by linoleic acid ethyl ester (11.05%), ethyl oleate, and hexadecanoic acid ethyl ester. Phenolic and aromatic compounds accounted for 14.24%, including (E)-4-(3-hydroxyprop-1-en-1-yl)-2-methoxyphenol and flamenol. Quantitative HPLC revealed hesperetin (69.72 µg/mL) and fisetin (12.32 µg/mL) as predominant in the 50% aqueous ethanol extract, and cinarin (6.28 µg/mL) in the ethyl acetate root extract. HPLC-MS identified 25 polyphenols, mainly isoflavonoids and flavones, with key markers songaricol, irilin B, tectorigenin, irisflavone A, and irizon B, some reported for the first time in Kazakhstan irises. Biological evaluation demonstrated potent activity: the 50% aqueous ethanol extract inhibited xylene-induced ear oedema in mice by 72.7% at 300 mg/kg, comparable to diclofenac (90.9%), without observable toxicity. These findings confirm I. songarica as a valuable source of bioactive polyphenols with anti-inflammatory potential. Full article

The rising demand for sustainable and circular approaches in the agro-industrial sector has generated interest in repurposing herbal tea residues as sources of high-value bioactive compounds. This work focusses on recovering phytochemicals from Rosa canina L. peel and seed dust (by-products of processing of herbal tea in filter tea bags) using green extraction techniques. Two environmentally friendly technologies were used: ultrasound-assisted extraction (UAE) with a sonotrode and subcritical fluid extraction (SBFE). The extracts were qualitatively profiled using (HR) LC-ESI-QToF-MS/MS and quantified using HPLC-PDA. Both by-products contained phenolic substances, including gallic acid derivatives, ellagic acid, and flavonoids such as quercetin and quercetin-3-O-glucoside (only in the peel). Additionally, Folin–Ciocalteu’s assay was used to determine Total Phenolic content (TP). The extraction efficiency was considered in terms of phenolic compound recovery and total phenolic content obtained under the respective experimental conditions. The maximum TP for SBFE was reported in samples extracted with ethanol–water (48:52) at 180 °C, producing 3876.67 GAE mg/L for peel and 1648.57 GAE mg/L for seeds. In the UAE, extraction with ethanol–water (48:52) for 10 min yielded the maximum TP of 2773.81 GAE mg/L for peel and 957.86 GAE mg/L for seeds. These findings highlight the potential of R. canina infusion by-products as long-term sources of bioactive compounds for use in nutraceutical, cosmetic, and pharmaceutical industries. Full article

This study provides a comparative evaluation of two Salvia species, the widely cultivated Salvia sclarea L. and the comparatively underexplored wild species Salvia pratensis L., integrating phytochemical profiling, chemical safety assessment, and biological activity investigation. Dried hydroethanolic extracts and essential oils obtained from aerial parts were analysed. HPLC–PDA analysis revealed distinct phenolic acid profiles, with S. sclarea characterized by higher levels of rosmarinic and protocatechuic acids, whereas S. pratensis contained greater amounts of hydroxycinnamic acids such as caffeic, p-coumaric, and ferulic acids. The total phenolic content was higher in S. pratensis (79.22 mg GAE/g dry extract) than in S. sclarea (52.50 mg GAE/g). GC–MS analysis showed that the essential oil of S. sclarea was dominated by oxygenated monoterpenes, mainly linalyl acetate and linalool, while S. pratensis exhibited a linalool-rich profile accompanied by sesquiterpene derivatives. Chemical safety assessment indicated minimal contamination, with pesticide residues detected only in S. sclarea at levels below regulatory limits and low concentrations of cadmium and lead in both species. The extracts showed strong antioxidant activity (DPPH IC₅₀ values of 6.67 µg/mL for S. sclarea and 3.16 µg/mL for S. pratensis) and moderate broad-spectrum antimicrobial activity (MIC 312.5–2500 µg/mL). In vitro assays on HEK 293 and HaCaT cells confirmed low cytotoxicity, with no evidence of membrane damage or pro-inflammatory effects. Overall, the results highlight the significant bioactive potential of the less studied S. pratensis, demonstrating that this wild species represents a promising alternative source of natural antioxidant and antimicrobial compounds comparable to the widely cultivated S. sclarea. Full article