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This study established a simple, rapid, and sensitive ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for the simultaneous quantification of contezolid and its major metabolite M2 in human plasma and cerebrospinal fluid (CSF). We used isotope-labeled internal standards (IS) along with protein precipitation to efficiently eliminate impurities. The ion transition was m/z 409.1 → 269.1 for contezolid, m/z 414.1 → 269.0 for contezolid-d5, m/z 445.1 → 385.1 for M2, and m/z 450.2 → 390.1 for M2-d5. The flow rate was 0.4 mL/min with a run time of 2.5 min. Validation results demonstrated that the calibration range was 10–5000 ng/mL for contezolid, and 5–2500 ng/mL for M2 in plasma and CSF. Both analytes exhibit excellent linear relationships (all R2 ≥ 0.99). Intra- and inter-batch accuracy and precision for all quality control (QC) levels were within ± 15%, including the lower limit of quantification (LLOQ). The recoveries of contezolid and M2 were 92.60–100.4% and 92.26–99.61% in plasma and 93.33–99.19% and 91.31–98.21% in CSF, respectively. The matrix effects for contezolid and M2 were 97.20–110.3% and 98.27–105.47% in plasma and 105.25–112.99% and 103.35–109.84% in CSF, respectively. The coefficients of variation (CV%) were ≤ 12.58%. Conteozolid and M2 remained stable in plasma and CSF. This study has established a simple and accurate method for determining contezolid and M2 in human plasma and CSF, making it a valuable tool for monitoring the efficacy of central nervous system (CNS) drugs. [ABSTRACT FROM AUTHOR]

Solar ultraviolet (UV)‐induced DNA lesions are associated with skin carcinogenesis. The detection of these DNA lesions is important to understand their genotoxicity and health effects. However, sunlight exposure‐relevant DNA damage measurement is still a challenge. Here, we summarize our recent progresses on the related analytical techniques, including synthesis of dimeric lesions, the optimization of procedures for ultrahigh performance liquid chromatography–coupled mass spectrometry (UHPLC‐MS/MS), and the maturation of anti‐T(6‐4)T photoproduct antibodies and their potential applications for immunoassay. [ABSTRACT FROM AUTHOR]

Solar ultraviolet (UV)‐induced DNA lesions are associated with skin carcinogenesis. The detection of these DNA lesions is important to understand their genotoxicity and health effects. However, sunlight exposure‐relevant DNA damage measurement is still a challenge. Here, we summarize our recent progresses on the related analytical techniques, including synthesis of dimeric lesions, the optimization of procedures for ultrahigh performance liquid chromatography–coupled mass spectrometry (UHPLC‐MS/MS), and the maturation of anti‐T(6‐4)T photoproduct antibodies and their potential applications for immunoassay. [ABSTRACT FROM AUTHOR]

The entity of DNA N6-methyladenine (6mA) in mammals remains elusive and subsequently its roles in diseases are poorly understood. Here we exploited a bacterial DNA contamination-free and ultrasensitive UHPLC-MS/MS assay to reassess DNA 6mA in human glioblastomas and unveiled that DNA 6mA (~0.08 ppm) is extremely rare. By the use of two independent heavy stable isotope-labeling strategies, we further prove that the observed 6mA is solely generated by DNA polymerase-mediated misinocorporation. In vitro experiments point toward that the generation of misincorporated DNA 6mA is associated with the cellular stresses-caused release of RNA N6-methyladenine (m6A) nucleoside, which is profoundly inhibited by hypoxia milieu. Consistently, compared with normal brain tissues, DNA 6mA decreases in hypoxic human gliomas. Our data also strongly support that rare DNA 6mA rather than relatively abundant DNA 5-methylcytosine and 5-hydroxymethylcytosine is a hallmark of poor prognosis of IDH1/2 mutation-absent glioblastoma patients, reflecting the incidence of cytotoxic stresses and subsequent release of m6A nucleoside. The released m6A nucleoside may selectively preserve a subset of the glioblastoma cells and stimulate their stemness and proliferation. Noteworthily, demethylation-inhibiting IDH1 mutation increases the DNA 6mA content in human gliomas, but the depletion of the demethylase candidate ALKBH1 fails to do so, together suggesting the presence of other unknown 6mA demethylase for erasing misincorporated DNA 6mA. This is the first report on the identification of the misincorporated 6mA together with its origin and roles in diseases. [ABSTRACT FROM AUTHOR]

Environmental exposure would cause DNA damage and epigenetic modification changes, potentially resulting in physiological dysfunction, thereby triggering diseases and even cancer. DNA damage and epigenetic modifications are thus promising biomarkers for environmental exposures and disease states. Benefiting from its high sensitivity and accuracy, high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) is considered the "gold standard technique" for investigating epigenetic DNA modifications. This review summarizes the recent advancements of UHPLC-MS/MS-based technologies for DNA damage and epigenetic modifications analysis, mainly focusing on the innovative methods developed for UHPLC-MS/MS-related pretreatment technologies containing efficient genomic DNA digestion and effective removal of the inorganic salt matrix, and the new strategies for improving detection sensitivity of liquid chromatography-mass spectrometry. Moreover, we also summarized the novel hyphenated techniques of the advanced UHPLC-MS/MS coupled with other separation and analysis methods for the measurement of DNA damage and epigenetic modification changes in special regions and fragments of chromosomes. • Accelerated genomic DNA digestion strategies for LC-MS analysis. • Matrix desalination and salt interference mitigation. • New strategies for improving detection sensitivity of LC-MS. • Applications of DNA damage and epigenetic modification analysis via LC-MS and complementary techniques. [ABSTRACT FROM AUTHOR]

This article explores the relationship between the fungus Ustilaginoidea virens and the WRKY transcription factor OsWRKY31 in rice plants. The researchers discovered that a specific protein, UvSec117, is a key factor in the virulence of U. virens and interacts with OsWRKY31. They also found that OsWRKY31 positively regulates rice's resistance to various diseases. Through further experiments, they identified the target genes of OsWRKY31, which are involved in induced systemic resistance and the salicylic acid/jasmonic acid signaling pathways. This research provides valuable insights into the molecular mechanisms of rice's resistance to grain-infecting pathogens and has implications for breeding disease-resistant rice varieties. [Extracted from the article]

Bisphenol F (BPF) is a major alternative to bisphenol (BPA) and has been widely used. Although BPA exposure is known to generate various toxic effects, toxicity of BPF remains under-explored. A comprehensive method involving mass spectrometry (MS)-based global lipidomics and metabolomics, and matrix-assisted laser desorption/ionization-mass spectrometry (MALDI)- MS imaging (MSI) was used to study toxic effects of BPF and the underlying mechanisms on tumor metastasis-related tissues (liver and kidney) in breast cancer xenografts. Our results demonstrated that BPF exposure disturbed the metabolome and lipidome of liver and kidney. Exposure induced reprogramming of the glutathione (GSH) biosynthesis and glycolytic metabolism by activating glycine, serine, cysteine, glutamine, lactate and pyruvate in liver and kidney tissues. It also perturbed the biosynthesis and degradation of glycerophospholipids (GPs) and glycerolipids (GLs), resulting in abnormality of membrane homeostasis and cellular functions in kidney tissues. Moreover, spatial distribution and profile of metabolites changed across renal cortex and medulla regions after BPF treatment. Levels of phosphatidylethanolamines (PE) and triacylglycerols (TAG) increased in renal medulla and pelvis, while the levels of phosphatidylcholines (PC) and phosphatidylinositols (PI) increased in cortex and pelvis. These observations offer a deeper understanding of critical role of metabolites and lipid reprogramming in BPF-induced biological effects. [ABSTRACT FROM AUTHOR]

Long-term exposures to environmental toxicants are causative factors for human health effects. DNA damages are of great interest in that they can serve as the potential biomarkers of environmental exposure and mutations. Qualitative and quantitative detection of their level in genomic DNA, are helpful for the understanding of the generation, transformation, repair and fate of DNA damages and for elucidating the mechanism underlying carcinogenesis. Methodologies based on liquid chromatography-mass spectrometry (LC-MS) have shown unique advantages in accurate and highly sensitive analysis of these structurally modified DNA damages. The present review summarizes our current knowledge on LC-MS analysis of genetic DNA damages, with special emphasis on structurally chemical DNA modifications induced by environmental exposure. Advances in LC-MS analysis and monitoring of oxidative DNA lesion, DNA adducts induced by heavy metals, persistent organic pollutants, bisphenols, fine particulate matters, aldehydes and halobenzoquinones are comprehensively summarized. • Highly sensitive LC-MS methods for DNA damages analysis. • Oxidative DNA damages, DNA adducts induced by environmental exposure. • DNA modifications caused by new emerging contaminants including PM2.5 and BPA. • Advances in DNA damage monitoring and adductomics benefited from LC-MS/MS. [ABSTRACT FROM AUTHOR]

Abstract Four DNA bases are essential biochemical elements for coding heritable life blueprint. Meanwhile, organisms utilize modifying enzymes to post chemical groups on DNA bases to regulate gene expression, chromosome structures, and transposase activity, and to defense pathogens. The researches on the biochemical modifications of DNA bases are at the cutting edge of life sciences and chemical biology. Up to date, along with fantastic progresses, liquid chromatography hyphenated with mass spectrometry (LC-MS)-based technologies become popular and powerful tools for chemical structure-dependent identification and quantification of diverse DNA modifications. Here we provided comprehensive review on LC-MS-based technologies and related pretreatment technologies for profiling of epigenetic DNA modifications, mainly focusing on innovative technologies developed for desalting, DNA digestion, and ionization for mass spectrometry detection. We also summarized the epigenetic progresses, which benefited from the development of the LC-MS methodologies and technologies, on the studies of specific DNA modifications, in particular, 5-hydroxymethylcytosine and 5-formylcytosine. Highlights • LC-MS/MS methods for detection of epigenetic DNA modifications. • DNA hydrolysis for generation of LC-MS detectable mononucleosides. • Chemical approaches for enhancing LC-MS signal. • Progresses on epigenetic DNA modifications benefited from LC-MS/MS analysis. [ABSTRACT FROM AUTHOR]

DNA N6-methyladenine modification (6mA) is a predominant epigenetic mark in prokaryotes but rarely present in multicellular metazoa. The analytical technologies have been developed for sensitive detection of 6mA, including ultra-high performance liquid chromatography coupled with mass spectrometry (UHPLC-MS/MS) and single molecule real-time sequencing (SMRTseq). However, it remains challenging to detect 6mA at global level and/or in the context of sequence in multicellular metazoa (including mammals). This mini-review brings insights into current dilemma and potential solutions for the identification and quantifications of 6mA in mammals. [ABSTRACT FROM AUTHOR]

• A recyclable and reusable multi-enzymes cascade (SNSC-MNPs) for enzymatic digestion of genomic DNA was constructed. • SNSC-MNPs cascade enable fast digestion of genomic DNA into mono-nucleosides within 30 min. • Coupling SNSC-MNPs with HPLC-MS/MS, a sensitive and accurate method for quantitation of 5-hydroxymethylcytosine (5hmC) was developed. • TCBQ exposure induces elevated 5hmC levels in the genome of T24 cells. Long-term exposure to halobenzoquinones (HBQs) can induce genomic damages and abnormal epigenetic modifications. High-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) has shown unique advantages in identification and sensitive analysis of these structurally modified DNA lesions. Prior to MS analysis, genomic DNA needs to be fully digested into mono-nucleosides. Here, we prepared Supernuclease (SN)-, snake venom phosphodiesterase (SVP)- and calf intestinal alkaline phosphatase (CIP)- individually immobilized magnetic nanoparticles (MNPs), and combined them according to certain formula to construct a recyclable SN-SVP-CIP magnetic nanoparticles (SNSC-MNPs) cascade for rapid and efficient DNA digestion. The SNSC-MNPs cascade can fully digest genomic DNA into mono-nucleosides within 30 min. The SNSC-MNPs cascade coupled with HPLC-MS/MS method can accurately and sensitively detect 5-hydroxymethylcytosine (5hmC) changes in genome of human bladder cancer T24 cells induced by tetrachlorobenzoquinone. The immobilization of enzymes on MNPs can enhance the stability and enzymatic activity of the three enzymes, which guarantees the reusability and longtime preservation of the cascades. The relative digestive efficiencies are among 86% -106% up to ten times of reuse. The newly synthesized SNSC-MNPs cascade coupled with HPLC-MS/MS method is promising for fast identification and analysis of structural modifications in genomic DNA. [ABSTRACT FROM AUTHOR]