Search Results (24)

Background: Although B vitamins are implicated in cardiovascular regulation, the associations between serum thiamine (vitamin B1) and blood pressure (BP) remain unclear, particularly among women who are at high risk for hypertension-related complications. This study aimed to investigate relationships between serum thiamine levels and BP outcomes among middle-aged and elderly women in eastern China. Methods: A community-based cross-sectional study was conducted among 2015 women aged 45–69 years in Zhejiang Province, China. Serum thiamine levels were quantified using liquid chromatography tandem mass spectrometry (LC-MS/MS). Hypertension was defined as measured BP ≥ 140/90 mmHg, or current use of antihypertensive medications. Multivariate logistic and linear regression models were used to assess associations of thiamine with hypertension prevalence and BP levels, respectively. Dose–response relationships were evaluated using restricted cubic splines (RCSs). Results: Higher thiamine levels were significantly associated with reduced hypertension prevalence (adjusted OR per SD increase: 0.87; 95%CI: 0.77, 0.97), with RCSs confirming linear dose–response (p-overall < 0.05, p-nonlinearity > 0.05). Compared with the lowest tertile, participants in the highest thiamine tertile had a 25% lower hypertension risk. Thiamine levels also showed negative associations with systolic BP (adjusted coef: −1.51 mmHg per SD; 95% CI: −2.33, −0.68), with the participants in the highest tertile showing a 3.94 mmHg reduction (95%CI: −5.97, −1.92). No significant relationship was found for diastolic BP. Conclusions: Serum thiamine is inversely associated with both hypertension prevalence and systolic BP in middle-aged and elderly women. This study supports the potential of serum thiamine as a modifiable biomarker in hypertension prevention strategies, particularly among aging women. Full article

Background/Objectives: Oxazolidinones are novel antimicrobial agents used to combat bacterial infections, particularly multidrug-resistant strains. However, the synthesis of oxazolidinone derivatives, such as linezolid, often involves the use of 3,4-difluoronitrobenzene (DFNB) as an initiator. Despite its effectiveness, residual DFNB in drug products raises significant health concerns due to its structural similarity to toxic and carcinogenic nitrobenzenes. This contamination is particularly concerning in pharmaceutical formulations, where it poses potential patient safety hazards. Therefore, strict concentration limits for this impurity are necessary. Methods: To ensure tight control of DFNB concentrations, this study established an 8.3 µg/g target limit. An advanced high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed to overcome current limitations in detecting trace DFNB. Under negative atmospheric pressure chemical ionization (APCI) conditions, DFNB exhibited characteristic ion formations, including [M]^•− through electron capture and [M − F + O]⁻ via substitution reactions. The quantitative method utilizes MS/MS ion transitions of the substitution product while optimizing chromatographic and spectrometric parameters to enhance both sensitivity and specificity. Conclusions: Validation tests confirm the efficiency, precision, and accuracy of this method, with a low limit of quantification (LOQ) of 5 ng/mL (0.83 µg/g). This technique enables accurate detection and quantification of DFNB in linezolid active pharmaceutical ingredient (API) and various formulations, providing a reliable tool for quality control. This method ensures the safe use of linezolid by effectively monitoring and minimizing the risks associated with DFNB contamination. Full article

Apple pomace, a by-product of apple juice production, is typically discarded as waste. Recent approaches have focused on utilizing apple pomace by extracting beneficial bioactive compounds, such as antioxidant phenolic compounds (PCs). Before these PC-rich extracts can be used in food products, they must undergo food preservation and processing methods. However, the effects of these processes on the composition, stability, and properties of the PC remain insufficiently understood. The present study aimed at investigating the effects of a thermal treatment (TT), a high-pressure thermal treatment (HPTT), and a pulsed electric field treatment (PEF) on the composition and antioxidant activity of PC-rich apple pomace extracts (APEs). Major PCs, including phloridzin, chlorogenic acid, and epicatechin, as well as minor compounds, were identified by liquid chromatography–tandem mass spectrometry (LC-MS/MS) and high-performance thin-layer chromatography (HPTLC). As a stability indicative property, the antioxidant activity was analyzed by a Trolox equivalent antioxidant capacity assay (TEAC), electron paramagnetic resonance spectroscopy, and the Folin–Ciocalteu reagent assay. The results showed that TT at 80 °C increased phloridzin content, likely due to the hydrolysis of bound forms, while higher temperatures and HPTT resulted in a substantial PC conversion. The PEF treatment also caused notable PC conversion, but generally, it had a milder effect compared to TT and HPTT. Hence, low temperatures with and without high pressure and PEF seem to be the most promising treatments for preserving the highest content of major PC in APE. Antioxidant activity varied among the analytical methods, with HPTT showing minor changes despite PC loss compared to the untreated APE. This suggests that other antioxidant compounds in the extracts may contribute to the overall antioxidant activity. This study demonstrates that apple pomace contains valuable PC, highlighting its potential as a health-promoting food additive and the impact of conventional preservation and processing methods on PC stability. Full article

Bcl-2-associated athanogene 3 (BAG3) plays an important function in cellular protein quality control (PQC) maintaining proteome stability. Mutations in the BAG3 gene result in cardiomyopathies. Due to its roles in cardiomyopathies and the complexity of BAG3–protein interactions, it is important to understand these protein interactions given the importance of the multifunctional cochaperone BAG3 in cardiomyocytes, using an in vitro cardiomyocyte model. The experimental assay was conducted using high pressure liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) in the human AC16 cardiomyocyte cell line with BioID technology. Proteins with BAG3-interaction were identified in all the 28 hallmark gene sets enriched in idiopathic cardiomyopathies and/or ischemic disease. Among the 24 hallmark gene sets enriched in both idiopathic cardiomyopathies and ischemic disease, 15 gene sets had at least 3 proteins with BAG3-interaction. This study highlights BAG3 protein interactions, unveiling the key gene sets affected in cardiomyopathies, which help to explain the molecular mechanisms of the cardioprotective effects of BAG3. In addition, this study also highlighted the complexity of proteins with BAG3 interactions, implying unwanted effects of BAG3. Full article

The study explored proteomics to better understand the relationship between type 2 diabetes (T2DM) and hypertension (HT) in Thai adults, using shotgun proteomics and bioinformatics analysis. Plasma samples were taken from 61 subjects: 14 healthy subjects (mean age = 40.85 ± 7.12), 13 with T2DM (mean age = 57.38 ± 6.03), 16 with HT (mean age = 66.87 ± 10.09), and 18 with coexisting T2DM/HT (mean age = 58.22 ± 10.65). Proteins were identified using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Protein–protein interactions were analyzed using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) version 11.5. We identified six unique proteins in T2DM patients, including translationally controlled 1 (TPT1) and nibrin (NBN), which are associated with the DNA damage response. In HT patients, seven unique proteins were identified, among them long-chain fatty acid-CoA ligase (ASCL), which functions in the stimulation of triacylglycerol and cholesterol synthesis, and NADPH oxidase activator 1 (NOXA1), which is involved in high blood pressure via angiotensin II-induced reactive oxygen species (ROS)-generating systems. In coexisting T2DM/HT patients, six unique proteins were identified, of which two—microtubule-associated protein 1A (MAP1A)—might be involved in dementia via RhoB-p53 and diacylglycerol kinase beta (DGKB), associated with lipid metabolism. This study identified new candidate proteins that are possibly involved in the pathology of these diseases. Full article

Propolis has gained popularity in recent years as a potential preventive and therapeutic agent due to its numerous health benefits, which include immune system boosting, blood pressure lowering, allergy treatment, and skin disease treatment. The pharmacological activity of propolis is primarily attributed to phenolics and their interactions with other compounds. Given that phenols account for most of propolis’s biological activity, various extraction methods are being developed. The resin–wax composition of the propolis matrix necessitates the development of an extraction procedure capable of breaking matrix–phenol bonds while maintaining phenol stability. Therefore, the aim of this study was to assess the stability of two major groups of phenolic compounds, flavonoids and phenolic acids, in propolis methanol/water 50/50 (v/v) extracts obtained after ultrasound-assisted extraction (USE) under different extraction parameters (extraction time and pH) and heat reflux extraction (HRE). The methodology involved varying the USE parameters, including extraction time (5, 10, and 15 min) and pH (2 and 7), followed by analysis using liquid chromatography–tandem mass spectrometry (LC-MS/MS) to quantify phenolic recoveries. Results revealed that benzoic acid and chlorogenic acid derivatives demonstrated excellent stability across all ultrasound extraction procedures. The recoveries of flavonoids were highly diverse, with luteolin, quercitrin, and hesperetin being the most stable. Overall, neutral pH improved flavonoid recovery, whereas phenolic acids remained more stable at pH = 2. The most important optimization parameter was USE time, and it was discovered that 15 min of ultrasound resulted in the best recoveries for most of the phenols tested, implying that phenols bind strongly to the propolis matrix and require ultrasound to break the bond. However, the high variability in phenol extraction and recovery after spiking the propolis sample shows that no single extraction method can produce the highest yield of all phenols tested. As a result, when working with a complex matrix like propolis, the extraction techniques and procedures for each phenol need to be optimized. Full article

Protein hydrolysates from sea cucumber (Apostichopus japonicus) gonads are rich in active materials with remarkable angiotensin-converting enzyme (ACE) inhibitory activity. Alcalase was used to hydrolyze sea cucumber gonads, and the hydrolysate was separated by the ultrafiltration membrane to produce a low-molecular-weight peptide component (less than 3 kDa) with good ACE inhibitory activity. The peptide component (less than 3 kDa) was isolated and purified using a combination method of ACE gel affinity chromatography and reverse high-performance liquid chromatography. The purified fractions were identified by liquid chromatography–tandem mass spectrometry (LC–MS/MS), and the resulting products were filtered using structure-based virtual screening (SBVS) to obtain 20 peptides. Of those, three noncompetitive inhibitory peptides (DDQIHIF with an IC₅₀ value of 333.5 μmol·L⁻¹, HDWWKER with an IC₅₀ value of 583.6 μmol·L⁻¹, and THDWWKER with an IC₅₀ value of 1291.8 μmol·L⁻¹) were further investigated based on their favorable pharmacochemical properties and ACE inhibitory activity. Molecular docking studies indicated that the three peptides were entirely enclosed within the ACE protein cavity, improving the overall stability of the complex through interaction forces with the ACE active site. The total free binding energies (ΔG_total) for DDQIHIF, HDWWKER, and THDWWKER were −21.9 Kcal·mol⁻¹, −71.6 Kcal·mol⁻¹, and −69.1 Kcal·mol⁻¹, respectively. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that HDWWKER could significantly decrease the systolic blood pressure (SBP) of SHRs after intravenous administration. The results showed that based on the better antihypertensive activity of the peptide in SHRs, the feasibility of targeted affinity purification and computer-aided drug discovery (CADD) for the efficient screening and preparation of ACE inhibitory peptide was verified, which provided a new idea of modern drug development method for clinical use. Full article

N-nitroso-N-methyl-4-aminobutyric acid (NMBA) is the third N-nitrosamine impurity found in sartans. Herein, a sensitive and stable LC-MS/MS method with multiple reactions monitoring mode has been developed for the quantitative determination of NMBA in four sartan substances. The effective separation of NMBA and sartan substances was achieved on a C18 column under gradient elution conditions. The mass spectrometry method of the atmospheric pressure chemical ionization source and internal standard method was selected as the quantitative analysis method of NMBA. Then, this proposed LC-MS/MS analysis method was validated in terms of specificity, sensitivity, linearity, accuracy, precision and stability. Good linearity with correlation coefficient over 0.99 was obtained at the NMBA concentration of 3–45 ng/mL, and the limit of quantification was 3 ng/mL. Additionally, the recoveries of NMBA in four sartan substances ranged from 89.9% to 115.7%. The intra-day and inter-day relative standard deviation values were less than 5.0%. In conclusion, this developed determination method for NMBA through liquid chromatography–tandem mass spectrometry showed the characteristics of good sensitivity, high accuracy and precision, which will be of great help for the quantitative analysis of NMBA in sartan products. Full article

Cyanotoxins are toxic metabolites produced by most cyanobacteria. In recent years, the occurrence of cyanobacterial blooms in aquatic ecosystems has temporally and spatially increased because of nutrient oversupply caused by human and also by climatic changes. This increase has a negative impact on water quality, ecosystem integrity, and human health. Cyanotoxins constitute a group of compounds with diverse physicochemical properties and their presence in drinkable, fishable, and recreational water is the main health-damaging cause. They are also able to bioaccumulate in plants and vegetables irrigated with contaminated water. Research on the development of suitable analytical methods is needed to establish early-warning strategies for the improved protectionof humans and ecosystems health. Liquid chromatography coupled with mass spectrometry (LC-MS) has been the preferred option for the control of these compounds, mainly using reverse-phase mode or hydrophilic interaction liquid chromatography (HILIC) in order to separate multiclass cyanotoxins of varying polarity, which cannot be handled by the commonly used reverse phase columns. In this work, we propose the use of capillary electrophoresis (CE) coupled with tandem mass spectrometry using triple quadrupole and positive electrospray ionization (CE-(ESI)-MS/MS) to determine a mixture of cyanotoxins with different polarity. CE is an advantageous alternative to LC given its short analysis times, high resolution, low sample and reagent volumes, and the use of silica capillaries and buffers as separation media, resulting in lower cost and low environmental impact. Moreover, CE allows the analysis of molecules hardly affordable by LC, such as polar and very similar compounds (e.g., isomers). The method is designed for the simultaneous determination of eight cyanotoxins belonging to three different classes: cyclic peptides (microcystin-LR, microcystin-RR, and nodularin), alkaloids (cylindrospermopsin, anatoxin-a), and three non-protein amino acids isomers (β-methylamino-L-alanine, 2,4-diaminobutyric acid, and N-(2-aminoethyl) glycine). Separation was achieved using an acidic background electrolyte (BGE) consisting in 2 M of formic acid (FA) and 20% acetonitrile in water. The proper separation and resolution of the three non-protein amino acid isomers was one of the main challenges of the method. This was overcome by applying a voltage of 30 kV in a 90 cm length capillary at 20 °C. Parameters affecting MS detection and the sheath–liquid interface were also studied. Finally, the fixed values were: a sheath gas flow rate of 5 L/min at 195 °C; sheath–liquid consists of MeOH/H2O/FA (50:49.95:0.05 v/v/v), a flow rate of 15 μL/min; and a nozzle voltage of 2000 V; N₂ dry gas rate of 11 L/min at 150 °C; a nebulizer pressure of 10 psi; and a capillary voltage of 2000 V. Online pre-concentration approaches were tested in order to achieve higher sensitivity, obtaining a enrichment factor of 4 with a mixed technique of pH-junction and Field Amplied Sample Stacking (FASS). Full article

In this study, a method of Bacillus subtilis spore inactivation under high pressure (P, 200 MPa) combined with moderate temperature (T, 80 °C) and the addition of antimicrobial peptide LK (10² μg/mL) was investigated. Spores presented cortex hydrolysis and inner membrane (IM) damage with an 8.16 log reduction in response to treatment with PT-LK, as observed by phase-contrast and inverted fluorescence microscopy and flow cytometry (FCM) analysis. Furthermore, a tandem mass tag (TMT) quantitative proteomics approach was utilized because Liquid chromatography-tandem mass spectrometry (LC–MS/MS) analysis data were used. After treatment with PT-LK, 17,017 polypeptides and 3166 proteins were detected from B. subtilis spores. Among them, 78 proteins showed significant differences in abundance between the PT-LK-treated and control groups, with 49 proteins being upregulated and 29 proteins being downregulated in the PT-LK-treated group. Genetic information processing, metabolism, cellular process, and environmental information processing were the main mechanisms of PT-LK-mediated spore inactivation. Full article

Golden Retrievers may suffer from Pnpl1-related inherited ichthyosis. Our study shows that in the stratum corneum (SC) of ichthyotic dogs, linoleic acid (LA) is also present in the form of 9-keto-octadecadienoic acid (9-KODE) instead of the acylacid form as in normal dogs. The fatty acids purified from SC strips (LA, acylacids) were characterized by liquid chromatography-tandem mass spectrometry (LC-MS) and atmospheric pressure chemical ionization (APCI). Electrospray ionization (ESI) and MS2(MS/MS Tandem mass spectrum/spectra)/M3 (MS/MS/MS Tandem mass spectrum/spectra) fragmentation indicated the positions of the double bonds in 9-KODE. We showed that ichthyotic dogs have a threefold lower LA content in the form of acylacids. The MS2 fragmentation of acyl acids showed in some peaks the presenceof an ion at the m/z 279, instead of an ion at m/z 293 which is characteristic of LA. The detected variant was identified upon MS3 fragmentation as 9-keto-octadecadienoic acid (9-KODE), and the level of this keto-derivative was increased in ichthyotic dogs. We showed by the APCI that such keto forms of LA are produced from hydroperoxy-octadecadienoic acids (HpODE) upon dehydration. In conclusion, the free form of 9-KODE was detected in ichthyotic SC up to fivefold as compared to unaffected dogs, and analyses by HPLC (High performance liquid chromatography) and ESI-MS (Electrospray Ionization-Mass Spectrometry) indicated its production via dehydration of native 9-HpODE. Full article

Postmenopausal women are at higher risk of developing cardiovascular diseases due to changes in lipid profile and body fat, among others. The aim of this study was to evaluate the association of urinary tartaric acid, a biomarker of wine consumption, with anthropometric (weight, waist circumference, body mass index (BMI), and waist-to-height ratio), blood pressure, and biochemical variables (blood glucose and lipid profile) that may be affected during the menopausal transition. This sub-study of the PREDIMED (Prevención con Dieta Mediterránea) trial included a sample of 230 women aged 60–80 years with high cardiovascular risk at baseline. Urine samples were diluted and filtered, and tartaric acid was analyzed by liquid chromatography coupled to electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). Correlations between tartaric acid and the study variables were adjusted for age, education level, smoking status, physical activity, BMI, cholesterol-lowering, antihypertensive, and insulin treatment, total energy intake, and consumption of fruits, vegetables, and raisins. A strong association was observed between wine consumption and urinary tartaric acid (0.01 μg/mg (95% confidence interval (CI): 0.01, 0.01), p-value < 0.001). Total and low-density lipoprotein (LDL) cholesterol were inversely correlated with urinary tartaric acid (−3.13 μg/mg (−5.54, −0.71), p-value = 0.016 and −3.03 μg/mg (−5.62, −0.42), p-value = 0.027, respectively), whereas other biochemical and anthropometric variables were unrelated. The results suggest that wine consumption may have a positive effect on cardiovascular health in postmenopausal women, underpinning its nutraceutical properties. Full article

Idiopathic normal pressure hydrocephalus (iNPH) is a potentially reversible neurological disease, causing motor and cognitive dysfunction and dementia. iNPH and Alzheimer’s disease (AD) share similar molecular characteristics, including amyloid deposition, t-tau and p-tau dysregulation; however, the disease is under-diagnosed and under-treated. The aim was to identify a panel of sphingolipids and proteins in CSF to diagnose iNPH at onset compared to aged subjects with cognitive integrity (C) and AD patients by adopting multiple reaction monitoring mass spectrometry (MRM-MS) for sphingolipid quantitative assessment and advanced high-resolution liquid chromatography–tandem mass spectrometry (LC–MS/MS) for proteomic analysis. The results indicated that iNPH are characterized by an increase in very long chains Cer C22:0, Cer C24:0 and Cer C24:1 and of acute-phase proteins, immunoglobulins and complement component fragments. Proteins involved in synaptic signaling, axogenesis, including BACE1, APP, SEZ6L and SEZ6L2; secretory proteins (CHGA, SCG3 and VGF); glycosylation proteins (POMGNT1 and DAG1); and proteins involved in lipid metabolism (APOH and LCAT) were statistically lower in iNPH. In conclusion, at the disease onset, several factors contribute to maintaining cell homeostasis, and the protective role of very long chains sphingolipids counteract overexpression of amyloidogenic and neurotoxic proteins. Monitoring specific very long chain Cers will improve the early diagnosis and can promote patient follow-up. Full article

This study was aimed at comparatively analyzing the sterols, tocopherols and fatty acids from edible flesh and processing waste obtained from three shrimp species, utilizing rapid liquid chromatography (LC)-atmospheric-pressure chemical ionization (APCI)-tandem mass spectrometry (MS/MS) and gas chromatography-mass spectrometry (GC-MS). Results revealed the presence of significantly (p < 0.05) high proportions of health-beneficial omega-3 (n3) polyunsaturated fatty acids (PUFAs) in Argentine red shrimp (34.3% in waste and 38.2% in the flesh), compared to black tiger shrimp (16.5–24.2%) and whiteleg shrimp (13.2–22.6%). Among sterols, cholesterol was found most dominant, accounting in the range 349.4 (white shrimp flesh) to 559.3 µg/g fresh weight (FW) (black shrimp waste). Surprisingly, waste was found to contain a substantially higher amount of α-tocopherol, for instance, 21.7 µg/g FW in edible flesh and 35.3 µg/g FW in the waste of black tiger shrimp. The correlation analysis indicated that shrimp with low total contents of lipids might have higher proportions of health-beneficial long-chain (LC)-n3-PUFAs eicosapentaenoic (EPA) and docosahexaenoic acid (DHA). The fat quality indices, including the high ratios of hypocholesterolemic (h)/hypercholesterolemic (H) fatty acids, and lowest values of the atherogenic index (AI) and thrombogenic index (TI) indicated the health-beneficial potential associated with fat intake from red shrimp. Overall, a significant amount of health-beneficial compounds in edible flesh of studied shrimp confers its extraordinary nutritional benefits. Moreover, considering the richness of processing waste with these compounds, their valorization can be prompted. Full article

Folates are a group of B₉ vitamins playing an important role in many metabolic processes such as methylation reactions, nucleotide synthesis or oxidation and reduction processes. However, humans are not able to synthesize folates de novo and thus rely on external sources thereof. Baker’s yeast (Saccharomyces cerevisiae) has been shown to produce high amounts of this vitamin but extensive identification of its folate metabolism is still lacking. Therefore, we optimized and compared different sample preparation and purification procedures applying solid phase extraction (SPE). Strong anion exchange (SAX), C18 and hydrophilic–lipophilic-balanced (HLB) materials were tested for their applicability in future metabolomics studies. SAX turned out to be the preferred material for the quantitative purification of folates. Qualification of several folate vitamers was achieved by ultra-high pressure liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-Q-ToF-MS) measurements and quantification was performed by liquid chromatography tandem mass spectrometry (LC-MS/MS) applying stable isotope dilution assays (SIDAs). The oxidation product s-pyrazino-triazine (MeFox) was included into the SIDA method for total folate determination and validation. Applying the best protocol (SAX) in regard to folate recovery, we analyzed 32 different vitamers in different polyglutamate states up to nonaglutamates, of which we could further identify 26 vitamers based on tandem-MS (MS²) spectra. Total folate quantification revealed differences in formyl folate contents depending on the cartridge chemistry used for purification. These are supposedly a result of interconversion reactions occurring during sample preparation due to variation in pH adjustments for the different purification protocols. The occurrence of interconversion and oxidation reactions should be taken into consideration in sample preparation procedures for metabolomics analyses with a focus on folates. Full article