Search

A method for the determination of 14 human milk oligosaccharides (HMOs) in infant formula milk powder was established by hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS). The sample was deproteinized by ethanol precipitation and degreased by freezing prior to separation by HILIC. The analytes were detected by multiple reaction monitoring (MRM) and quantified by an external standard method. Under optimized conditions, all HMOs showed good linear relationships within their respective concentration ranges, with determination coefficients (R2) greater than 0.994. The average recovery rates from blank goat’s and cow’s milk powders at three spiked levels ranged from 90.3% to 109.2%, with relative standard deviation (RSD) in the range of 0.7%–9.7% (n = 6), and the limit of detection (LOD) between 0.002 and 0.048 μg/mL. The developed method is characterized by fast analysis speed, high sensitivity and good accuracy, and meets the requirements for the rapid detection of the 14 HMOs in infant formula milk powder.

Colorectal cancer and gastric cancer are the most prevalent gastrointestinal malignancies worldwide, and early detection of these cancers is crucial to reduce their incidence and mortality. RNA methylation plays an important regulatory role in a variety of physiological activities, and it has drawn great attention in recent years. Methylated adenosine (A) modifications such as N6-methyladenosine (m6A), N1-methyladenosine (m1A), 2′-O-methyladenosine (Am), N6,2′-O-dimethyladenosine (m6Am), and N6,N6-dimethyladenosine (m62A) are typical epigenetic markers of RNA, and they are closely correlated to various diseases including cancer. Serum is a valuable source of biofluid for biomarker discovery, and determination of these adenosine modifications in human serum is desirable since they are emerging biomarkers for detection of diseases. In this work, a targeted quantitative analysis method using hydrophilic interaction liquid chromatography–tandem mass spectrometry (HILIC-MS/MS) was developed and utilized to analyze these methylated adenosine modifications in serum samples. The concentration differences between the healthy volunteers and cancer patients were evaluated by Mann–Whitney test, and receiver operator characteristic (ROC) curve analysis was performed to access the potential of these nucleosides as biomarkers. We demonstrated the presence of the m6Am in human serum for the first time, and we successfully quantified the concentrations of A, m6A, m1A, and m6Am in serum samples from 99 healthy controls, 51 colorectal cancer patients, and 27 gastric cancer patients. We found that the levels of m6A and m6Am in serum were both increased in colorectal cancer or gastric cancer patients, compared to that in healthy controls. These results indicate that m6A and m6Am in serum may act as potential biomarkers for early detection and prognosis of colorectal cancer and gastric cancer. In addition, the present work will stimulate investigations on the effects of adenosine methylation on the initiation and progression of colorectal cancer and gastric cancer.

Hydrophilic iridoid glycosides and oligosaccharides are the major active ingredients of Radix Rehmanniae. Analysis of oligosaccharides is a challenging task because they are highly hydrophilic, with similar chemical structures and absence of chromophore and fluorophore groups. The difficulty for simultaneous analysis of iridoid glycosides and oligosaccharides in Radix Rehmanniae is increased due to the polarity difference between the two types of ingredients. Objective: To develop a method for simultaneous determination of iridoid glycosides (ajugol, catalpol) and oligosaccharides (sucrose, melibiose, raffinose, mannotriose and stachyose) in Radix Rehmanniae. Methodology: Microwave‐assisted extraction (MAE) was established to extract target analytes from Radix Rehmanniae samples using methanol–water (60:40, v/v) as the extraction solvent. Fast separation of seven analytes was achieved by hydrophilic interaction liquid chromatography (HILIC) using an Accucore‐150‐Amide‐HILIC column. Sensitive and selective detection of the analytes was performed by triple quadrupole tandem mass spectrometry (TQ‐MS/MS) using multiple reaction monitoring in positive electrospray ionisation mode. Results: Good linearities were achieved for all the analytes with the correlation coefficients above 0.9991. The precisions resulted in deviations of less than 5.0% and the recoveries ranged from 93.8% to 105.5%. The established method was successfully applied to the analysis of iridoid glycosides and oligosaccharides in 12 samples of crude and processed Radix Rehmanniae. Conclusion: A simple, rapid and sensitive method based on MAE combined with HILIC‐UHPLC‐TQ‐MS/MS was developed for simultaneous determination of iridoid glycosides and oligosaccharides in Radix Rehmanniae for the first time. The method exhibited excellent performance with simple sample preparation, short analysis time, high selectivity and sensitivity. A simple, rapid and sensitive method based on microwave‐assisted extraction (MAE) combined with hydrophilic interaction liquid chromatography ultra‐high‐performance liquid chromatography coupled to triple quadrupole tandem mass spectrometry (HILIC‐UHPLC‐TQ‐MS/MS) was developed for simultaneous determination of iridoid glycosides and oligosaccharides in Radix Rehmanniae for the first time. The developed method was successfully applied to determine two iridoid glycosides and five oligosaccharides in crude and processed Radix Rehmanniae. The method exhibited excellent performance with simple sample preparation, short analysis time, high selectivity and sensitivity. [ABSTRACT FROM AUTHOR]

Abstract 5-Methyl-2′-deoxycytidine (5-mdC), 5-hydroxymethyl-2′-deoxycytidine (5-hmdC), 5-methylcytidine (5-mrC) and 5-hydroxymethylcytidine (5-hmrC) are epigenetic marks of DNA and RNA, and aberrant levels of these modified nucleosides were found to be associated with various cancers. Urine is a preferred source of biological fluid for biomarker discovery because the sample collection process is not invasive to patients. Herein, we developed a novel malic acid-enhanced hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for sensitive and simultaneous quantification of the modified cytosine nucleosides in human urine. Malic acid markedly increased the detection sensitivities of all four cytosine nucleosides, with the limits of detection (LODs) for 5-mdC, 5-hmdC, 5-mrC and 5-hmrC being 0.025, 0.025, 0.025 and 0.050 fmol, respectively. By using this method, we demonstrated, for the first time, the presence of 5-hmrC in human urine, and we successfully quantified 5-mdC, 5-hmdC, 5-mrC and 5-hmrC in urine samples collected from 90 patients with colorectal cancer (CRC) and 90 healthy controls. We found that the levels of 5-mdC, 5-hmdC, 5-mrC and 5-hmrC in urine were all substantially decreased in CRC patients, suggesting that these modified nucleosides might have great potential to be noninvasive biomarkers for early detection and prognosis of CRC. Together, we established a novel and sensitive method for detecting 5-methylated and 5-hydroxymethylated cytosine nucleosides in human urine and the results from this study may stimulate future investigations about the regulatory roles of these cytosine derivatives in the initiation and development of CRC. Graphical abstract Image 1 Highlights • A novel method was reported for analysis of 5-methylated and 5-hydroxymethylated cytosine nucleosides in urine. • Enhancement of detection sensitivity was achieved with the assistance of malic acid. • Accurate quantification was realized by using stable isotope dilution method. • 5-hydroxymethylcytidine was firstly discovered in human urine. • The modified cytosine nucleosides are present in significantly lower levels in urine of colorectal cancer patients. [ABSTRACT FROM AUTHOR]

Inositol pyrophosphates regulate diverse physiological processes; to better understand their functional roles, assessing their tissue-specific distribution is important. Here, we profiled inositol pyrophosphate levels in mammalian organs using an originally designed liquid chromatography-mass spectrometry (LC-MS) protocol and discovered that the gastrointestinal tract (GIT) contained the highest levels of diphosphoinositol pentakisphosphate (IP 7 ) and its precursor inositol hexakisphosphate (IP 6 ). Although their absolute levels in the GIT are diet dependent, elevated IP 7 metabolism still exists under dietary regimens devoid of exogenous IP 7 . Of the major GIT cells, enteric neurons selectively express the IP 7 -synthesizing enzyme IP6K2. We found that IP6K2-knockout mice exhibited significantly impaired IP 7 metabolism in the various organs including the proximal GIT. In addition, our LC-MS analysis displayed that genetic ablation of IP6K2 significantly impaired IP 7 metabolism in the gut and duodenal muscularis externa containing myenteric plexus. Whole transcriptome analysis of duodenal muscularis externa further suggested that IP6K2 inhibition significantly altered expression levels of the gene sets associated with mature neurons, neural progenitor/stem cells, and glial cells, as well as of certain genes modulating neuronal differentiation and functioning, implying critical roles of the IP6K2-IP 7 axis in developmental and functional regulation of the enteric nervous system. These results collectively reveal an unexpected role of mammalian IP 7 -a highly active IP6K2-IP 7 pathway is conducive to the enteric nervous system., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The influence of sucrose combustion products on smoking and nicotine addiction is still controversial because the presence of the sucrose may be treated as a source of aldehydes and organic acids. In e-liquids used as refills for electronic cigarettes, which are made primarily of poly(propylene glycol), glycerine and ethanol, sucrose may be present at trace levels, and its impact on mainstream smoke formation, and hence on human health and smoking/nicotine addiction is unknown. An analytical method was developed where high-performance liquid chromatography in hydrophilic interaction liquid chromatography mode and tandem mass spectrometry were used for fast and simple determination of sucrose and other saccharides in e-liquids for electronic cigarettes. Minimal effort was required in the sample preparation step, and satisfactory results were obtained, and the sample matrix had an insignificant impact. The chromatographic separation was done using an Ascentis Express OH5 column (150 mm × 2.1 mm, 2.7 μm). The coefficients of variation for within-day precision for three concentrations were 2.4 %, 1.6 % and 2.3 %, and the between-day coefficients of variation for a single concentration were 2.1 %, 2.5 % and 1.7 % measured on the next 3 days. The detection limit was 0.73 μg/g, and the sucrose content in e-liquids ranged from 0.76 to 72.93 μg/g among 37 samples. Moreover, with the method presented it is possible to determine the presence of other saccharides such as fructose, glucose, maltose and lactose. However, only sucrose was found in all samples of e-liquids. The proposed method is rapid, simple and reliable in terms of high-performance liquid chromatography coupled with tandem mass spectrometry. [ABSTRACT FROM AUTHOR]

A rapid and simple microanalysis method using TopTip C18spin tips, which are new micropipette tip spin columns for solid-phase extraction, was developed and validated for quantitative determination of methamphetamine and amphetamine in human plasma. All the solid-phase extraction procedures, including conditioning, sample loading, washing, and elution, using the TopTip C18spin tips, were performed by centrifugation. The analytes were extracted within 7–8 min from 0.1 mL of plasma. The methanol-rich eluate was injected directly into a hydrophilic interaction liquid chromatography/tandem mass spectrometer in positive electrospray ionization mode with selected reaction monitoring. The analytes were separated on an Imtakt Unison UK-silica column using a gradient with methanol and 10 mmol/L ammonium acetate buffer as mobile phase at a flow rate of 0.5 mL/min. The recoveries of methamphetamine and amphetamine spiked into plasma were 82–95% and the limit of quantification of methamphetamine and amphetamine were 2.5 ng/mL of plasma. The calibration curves were linear over the concentration range of 2.5–500 ng/mL of plasma for both compounds. The intra-day and inter-day relative standard deviations were less than 7.5%. This fast and simple method is expected to be very useful for quantitative determination of methamphetamine and amphetamine in human plasma samples. [ABSTRACT FROM PUBLISHER]

Lowered plasma concentrations of the endogenous amino acid l-homoarginine have been recently identified as an independent risk factor for cardiovascular and all-cause mortality in patients referred for coronary angiography in the LURIC study. To support further investigations into this matter, we describe here a fast and easy LC–MS–MS method for the detection of l-homoarginine in human plasma. The sample preparation consisted only of the addition of the stable isotope-labeled internal standard d4- l-homoarginine and protein precipitation. The analytes were separated isocratically on an HILIC silica column. Detection took place by tandem mass spectrometry. The calibration function was linear in the range of 0.1–10 μmol L −1. The intra-day precision was better than 2 % RSD and the inter-day precision better than 4 % RSD in plasma. The accuracy was always better than 5 % deviation. The method was matrix independent owing to the usage of the analogous stable isotope-labeled internal standard. [ABSTRACT FROM AUTHOR]

Monitoring carnitine and acetylcarnitine levels in biological fluids is a powerful tool for diagnostic studies. Research has recently shown that the analysis of carnitine and related compounds in clinical samples can be accomplished by different analytical approaches. Because of the polar and ionic nature of the analytes and matrix complexity, accurate quantitation is a highly challenging task. Thus, sample processing factors, preparation/cleanup procedures, and chromatographic/ionization/detection parameters were evaluated. On the basis of the results obtained, a rapid, selective, sensitive method based on hydrophilic interaction liquid chromatography-tandem mass spectrometry for the analysis of carnitine and acetylcarnitine in serum and urine samples is proposed. The matrix effect was assessed. The proposed approach was validated, the limits of detection were in the nanomolar range, and carnitine and acetylcarnitine levels were found within the micromolar range in both types of sample. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

Abstract: Carboplatin is a platinum agent that is used for treatment of non-small-cell lung cancer and ovarian cancer. A sensitive and selective analytical method for the quantification of carboplatin in human plasma ultrafiltrates using liquid chromatography–tandem mass spectrometry was developed. Human plasma ultrafiltrates were precipitated by acetonitrile containing carboplatin-d4 as an internal standard and were further diluted with acetonitrile. Chromatographic separation was performed on a Accucore HILIC (50mm×2.1mm i.d., 2.6μm) column using mobile phase (acetonitrile–water–acetic acid=90:10:0.1, v/v/v) at the flow rate of 0.2mL/min. Detection was performed on electrospray ionization triple quadrupole tandem mass spectrometer using low-energy collision induced dissociation (CID-MS/MS) analysis operating in the selected reaction monitoring (SRM) scan mode. The lower limit of quantification for carboplatin was 0.025μg/mL. This method covered a linearity range of 0.025–50μg/mL. The intra-day precision and inter-day precision (R.S.D.) ranged from 1.5 to 4.3%, and the accuracy (R.E.) was within ±2.9%. The present method was applied to a clinical pharmacokinetic study of carboplatin in a cancer patient. [Copyright &y& Elsevier]

Abstract: Oxaliplatin is a platinum agent that is used for treatment of colorectal cancer. A sensitive and selective hydrophilic interaction liquid chromatography–tandem mass spectrometric method for the quantification of oxaliplatin was developed. Human plasma ultrafiltrates were precipitated by acetonitrile containing carboplatin as an internal standard and further diluted with acetonitrile. Chromatographic separation of oxaliplatin and the internal standard was achieved with a column modified with phosphorylcholine and an isocratic mobile phase (acetonitrile/water/acetic acid=90:10:0.1, v/v/v) at the flow rate of 0.2mL/min. The lower limit of quantification for oxaliplatin was 25ng/mL. The linearity range of the method was from 25 to 5000ng/mL. The intra-day precision and inter-day precision (RSD) ranged from 0.8 to 6.1%, and the accuracy (RE) was within ±4.5%. The extraction recoveries from human plasma ultrafiltrates were 83.6–91.6%, and ion suppression caused by matrix components was 86.7–88.5% at three different levels, respectively. This method was applied to a clinical pharmacokinetic study of oxaliplatin in a cancer patient. The maximum concentration of colorectal cancer patient administered oxaliplatin was 1650ng/mL. [Copyright &y& Elsevier]