Scheduled Multiple Reaction Monitoring Research Articles

Rapid determination of 111 anti-infective drugs possibly added in cosmetics using high-performance liquid chromatography-tandem mass spectrometry with scheduled multiple reaction monitoring.

Illegal addition of anti-infective drugs to cosmetics at low concentrations has been found. The illicit addition of anti-infective drugs encompasses a wide variety of medications. The current sample purification methods are inadequate to detect all these compounds. A sensitive, wide-coverage, and weak-matrix-effect measurement method needs to be established to address this issue. Samples were extracted using acetonitrile, diluted 25 times, and then analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) to detect 111 anti-infective drugs. The method was validated and assessed for matrix effect before being applied to cosmetic products. The calibration curves for the analytes exhibited a strong correlation coefficient (r > 0.995). The limit of detection ranged from 0.006 to 0.6mg/kg. Matrix effects were significantly improved after a 25-fold dilution. The method was successfully applied to various cosmetics. Two of 82 samples tested contained lincomycin and miconazole, respectively. The developed method is quick and reliable to analyze anti-infective drugs in cosmetics, with potential for both qualitative and quantitative analyses. It is a valuable tool for cosmetic research and development, contributing to safer and more effective cosmetic products.

Disulfide-adducts with cysteine residues in human serum albumin prove exposure to malodorous mercaptans in vitro

Malodorants are mixtures containing mercaptans, which trigger the flight instinct upon exposure and might thus be deployed in military and civilian defense scenarios. Exposure to mercaptans might lead to unconsciousness, thus representing a possible threat for health. Therefore, we developed and validated a bioanalytical procedure for the simultaneous detection and identification of corresponding biomarkers for the verification of exposure to mercaptans. Disulfide-adducts of ethyl mercaptan (SEt), n-butyl mercaptan (SnBu), tert-butyl mercaptan (StBu) and iso-amyl mercaptan (SiAm) with cysteine (Cys) residues in human serum albumin (HSA) were formed by in vitro incubation of human plasma. After pronase-catalyzed proteolysis, reaction products were identified as adducts of the single amino acid Cys and the dipeptide cysteine-proline (Cys34Pro) detected by a sensitive μLC-ESI MS/MS method working in the scheduled multiple reaction monitoring (sMRM) mode. Dose-response studies showed linearity for the yield of Cys34Pro-adducts in the range from 6 nM to 300 μM of mercaptans in plasma and limits of identification (LOI) were in the range from 60 nM to 6 μM. Cys34-adducts showed stability for at least 6 days in plasma (37°C). The presented disulfide-biomarkers expand the spectrum for bioanalytical verification procedures and might be helpful to prove exposure to malodorants.

Ultra-high performance liquid chromatography-tandem mass spectrometry assay for simultaneous determination of 22 marker compounds in traditional herbal medicine Ojeoksan

Traditional herbal medicines (THMs) have long been in use worldwide and are considered safe for use as tonics or complementary treatments for many diseases. Advanced quality control methods for THMs are required in the regulatory framework of modern medicines. In this study, an ultra-high performance liquid chromatography-tandem mass spectrometry assay was established for the simultaneous determination of 22 marker compounds in Ojeoksan (OJS), which is composed of 15 herbal substances. All marker compounds were analyzed within 20 min and successfully identified via scheduled multiple reaction monitoring. The method validation revealed excellent performance characteristics of the method such as specificity, linearity, sensitivity, precision, and accuracy, demonstrating its suitability for intended use. The developed method was applied to samples of commercial OJS tablet and soft-extract dosage forms. The 14 marker compounds corresponding to 12 component herbal substances were determined in the samples; ephedirine, albiflorin, paeoniflorin, ferulic acid, hesperidine, neohesperidin, cinnamic acid, platycodin D, 6-gingerol, atractylenolide III, glycyrrhizin, honokiol, decursin, and magnolol. A fast and easy assay method with sufficient discrimination power was established. As a novel assay, this method can contribute to the quality control of OJS products.

Development and validation of an LC-MS/MS method for quantifying diamorphine and its major metabolites 6-monoacetylmorphine, morphine, morphine-3-glucuronide, and morphine-6-glucuronide in human plasma

Diamorphine, commonly known as heroin, is a semi-synthetic opioid analgesic. In the context of heroin-assisted treatment for opioid-dependent patients, diamorphine is mostly administered intravenously. However, recent attention has shifted towards intranasal administration as a better-tolerated alternative to the intravenous route. Here, we developed and validated a rapid bioanalytical method for the simultaneous quantification of diamorphine and its major metabolites 6-monoacetylmorphine, morphine, morphine-3-glucuronide, and morphine-6-glucuronide in human plasma using liquid chromatography-tandem mass spectrometry (LC-MS/MS).A straightforward protein precipitation extraction step was used for sample preparation. Chromatographic analyte separation was achieved using a Kinetex EVO C18 analytical column and a mobile phase gradient comprising an aqueous solution of ammonium hydrogen carbonate and methanol supplied with formic acid. Employing positive electrospray ionization and scheduled multiple reaction monitoring, we established a quantification range of 1–1,000 ng/mL for all analytes.Our validation results demonstrate a mean intra-assay accuracy of 91–106% and an intra-assay precision (CV) between 2 and 9% for all analytes and over three validation runs. The method exhibits a high extraction recovery (> 87%) and a negligible matrix effect (99–125%). Furthermore, no interferences with endogenous plasma compounds were detected. Lastly, we applied the method to assess the plasma concentrations of an opioid-dependent patient after the intranasal administration of diamorphine in a clinical study.In summary, we have successfully developed a rapid, highly reliable, and straightforward bioanalytical method for quantifying diamorphine and its metabolites in low amounts of clinical plasma samples.

Determination of 145 pharmaceuticals and personal care products in eleven categories in water by ultra-high performance liquid chromatography-triple quadrupole mass spectrometry

Pharmaceuticals and personal care products (PPCPs) are emerging contaminants frequently detected in aquatic environments at trace levels. These chemicals have diverse structures and physicochemical properties and includes pharmaceuticals like antibiotics, antihypertensive drugs, antiviral drugs, and psychotropic drugs that are widely used in large quantities worldwide. Considering the large number of pharmaceuticals currently in usage, it is crucial to establish a priority list of PPCPs that should be monitored and/or treated first. An accurate understanding of the occurrence and levels of PPCPs in aquatic environments is essential for providing objective materials for monitoring these emerging contaminants. Therefore, accurate, efficient, sensitive, and high-throughput screening techniques need to be established for determining and quantifying PPCPs. This study developed a method for the determination of 145 PPCPs (grouped into eleven categories: antibiotics, antihypertensive drugs, antidiabetic drugs, antiviral drugs, β-receptor agonists, nitroimidazoles, H2 receptor antagonists, psychotropic drugs, hypolipidemic drugs, non-steroidal anti-inflammatory drugs, and others) in water. The method was based on large volume direct injection without sample enrichment and cleanup and used ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS). Water samples were collected and filtered through a 0.22-μm regenerated cellulose (RC) filter membrane. Subsequently, Na2EDTA was added to the samples to adjust their pH to 6.0-8.0. Internal standards were mixed with the solutions, and because of the addition of Na2EDTA, the interference of metal ions could be eliminated in the determination of compounds, especially for tetracycline and quinolone antibiotics. Among the six filter membranes tested in this study (PES, PFTE-Q, PFTE, MCE, GHP, and RC), RC filter membranes were screened for water sample filtration. The UHPLC-MS/MS parameters were optimized by comparing the results of various mobile phases, as well as by establishing the best instrumental conditions. The 145 PPCPs were separated using an Phenomenex Kinetex C18 column (50 mm×3 mm, 2.6 μm) via gradient elution. The mobile phases were 0.1% (v/v) formic acid aqueous solution containing 5 mmol/L ammonium formate and acetonitrile for positive ion modes, 5 mmol/L aqueous solutions of ammonium formate and acetonitrile for negative ion modes. The samples were quantified using the scheduled multiple reaction monitoring (scheduled-MRM) mode with electrospray ionization in positive and negative ion modes. A standard internal calibration procedure was used to calculate contents of sample. The established method was systematically verified, and it demonstrated a good linear relationship. The average recoveries of the 145 PPCPs at the three spiked levels were in the range of 80.4%-128% with relative standard deviations (RSDs, n=6) of 0.6%-15.6%. The method detection limits (MDLs) ranged from 0.015 to 5.515 ng/L. Finally, the optimization method was applied to analyze the 145 PPCPs in 11 surface water samples and 6 drinking water samples. Overall, 93 (64%) out of the 145 analytes were detected. The total contents of the PPCPs in surface water samples ranged from 276.9 to 2705.7 ng/L. The detection frequencies of antidiabetic, antiviral, and psychotropic drugs were 100%. The total contents of the PPCPs in drinking water samples ranged from 140.5 to 211.5 ng/L, and antibiotics, antidiabetic drugs, and antiviral drugs comprised the largest proportion of analytes (by mass concentration) in drinking water samples. Our method exhibited high analytical speed and high sensitivity. It is thus suitable for the trace analysis and determination of the 145 PPCPs in environmental water and showed improved detection efficiency for PPCPs in water, indicating that it has a high potential for practical applications. This study can extend technical support for further pollution-level analysis of PPCPs in water and provide an objective basis for environmental management.

Quantitative analysis of tetrahydrocannabinol isomers and other toxicologically relevant drugs in blood.

A multi-analyte liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is described, involving the separation of delta-9-tetrahydrocannabinol (delta-9-THC) and delta-8-THC in addition to other commonly encountered drugs and metabolites. Briefly, sample preparation involved an alkaline liquid-liquid extraction (methyl tert-butyl ether) of blood (100 μl). The solvent layer was transferred, evaporated to dryness, reconstituted, and samples then separated on an Agilent Poroshell 120 EC-C18 100 Å (50 mm × 3.0mm, 2.7μm) analytical column using a multi-step gradient elution of 50 mM ammonium formate in water (pH3.5) and 0.1% formic acid in methanol over 14 min. A SCIEX Triple Quad 6500+ system operating in scheduled multiple reaction monitoring and positive electrospray ionization was used for detection. There were no interferences, and matrix effects were generally acceptable (±20% of neat response). Linearity was achieved within the calibration range, including methylamphetamine (MA) (10-1000 ng/ml), 3,4-methylenedioxy-N-methylamphetamine (MDMA) (10-1,000 ng/ml), cocaine (10-1000 ng/ml), and two THC isomers (1-100 ng/ml). Accuracies of MA, MDMA, cocaine, and two THC isomers were 3.6 to 8.9%, -1.2 to 4%, -5.3 to 5.8%, and -11 to 14%, respectively; while precision estimates of the same were 1.6 to 5.4%, 1.7 to 5.3%, 1.2 to 4.5%, and 2 to 10%, respectively. Autosamplerstability and dilution integrity were within acceptable limits, and no carryover was detected at the limit of detection. This validated LC-MS/MS method made the routine identification of both delta-9-THC and delta-8-THC in blood possible.

Development of targeted hydrophilic interaction liquid chromatography-tandem mass spectrometry method for acyl-Coenzyme A covering short- to long-chain species in a single analytical run

Acyl-CoAs play a significant role in numerous physiological and metabolic processes making it important to assess their concentration levels for evaluating metabolic health. Considering the important role of acyl-CoAs, it is crucial to develop an analytical method that can analyze these compounds. Due to the structural variations of acyl-CoAs, multiple analytical methods are often required for comprehensive analysis of these compounds, which increases complexity and the analysis time. In this study, we have developed a method using a zwitterionic HILIC column that enables the coverage of free CoA and short- to long-chain acyl-CoA species in one analytical run. Initially, we developed the method using an LC-QTOF instrument for the identification of acyl-CoA species and optimizing their chromatography. Later, a targeted HILIC-MS/MS method was created in scheduled multiple reaction monitoring mode using a QTRAP MS detector. The performance of the method was evaluated based on various parameters such as linearity, precision, recovery and matrix effect. This method was applied to identify the difference in acyl-CoA profiles in HepG2 cells cultured in different conditions. Our findings revealed an increase in levels of acetyl-CoA, medium- and long-chain acyl-CoA while a decrease in the profiles of free CoA in the starved state, indicating a clear alteration in the fatty acid oxidation process.

Global Analysis of 2-Hydroxy Fatty Acids by Gas Chromatography-Tandem Mass Spectrometry Reveals Species-Specific Enrichment in Echinoderms.

Abnormal levels of 2-hydroxy fatty acids (2-OH FAs) are characterized in multiple diseases, and their quantification in foodstuffs is critical to identify the sources of supplementation for potential treatment. However, due to the structural complexity and limited available standards, the comprehensive profiling of 2-OH FAs remains an ongoing challenge. Herein, an innovative approach based on gas chromatography-tandem mass spectrometry (GC-MS/MS) was developed to determine the full profile of these FA metabolites. MS and MS/MS spectra of the trimethylsilyl (TMS) derivatives of 2-OH fatty acid methyl esters (FAMEs) were collected for peak annotation by their signature fragmentation patterns. The structures were further confirmed by validated structure-dependent retention time (RT) prediction models, taking advantage of the correlation between the RT, carbon chain length, and double bond number from commercial standards and pseudostandards identified in the whole-brain samples from mice. An in-house database containing 50 saturated and monounsaturated 2-OH FAs was established, which is expandible when additional molecular species with different chain lengths and backbone structures are identified in the future. A quantitation method was then developed by scheduled multiple reaction monitoring (MRM) and applied to investigate the profiling of 2-OH FAs in echinoderms. Our results revealed that the levels of total 2-OH FAs in sea cucumber Apostichopus japonicas (8.40 ± 0.28 mg/g dry weight) and starfish Asterias amurensis (7.51 ± 0.18 mg/g dry weight) are much higher than that in sea urchin Mesocentrotus nudus (531 ± 108 μg/g dry weight). Moreover, 2-OH C24:1 is the predominant molecular species accounting for 67.9% of the total 2-OH FA in sea cucumber, while 2-OH C16:0 is the major molecular species in starfish. In conclusion, the current innovative GC-MS approach has successfully characterized distinct molecular species of 2-OH FAs that are highly present in sea cucumbers and starfish. Thus, these findings suggest the possibility of developing future feeding strategies for preventing and treating diseases associated with 2-OH FA deficiency.

Authenticity identification of animal species in characteristic milk by integration of shotgun proteomics and scheduled multiple reaction monitoring (MRM) based on tandem mass spectrometry

Milk is one of the oldest natural dairies with high value, which has different species including cow, camel, donkey, goat, sheep, buffalo, yak and et al. However, economically motivated adulteration of non-cow milk with cheaper cow milk occurs frequently. To develop a high-throughput approach for milk species authentication, integration of shotgun proteomics and scheduled multiple reaction monitoring (MRM) was developed. In total, 37 specific peptides were screened as unique to different species. Specific peptides processing stability was investigated under different treatment (heat, pressure, fermentation). Subsequently, four quantitative ion pairs of peptides from cow milk and six quantitative ion pairs of peptides from six non-cow milks were selected for the adulteration quantitative analysis. The method is capable of detection adulteration in the range of 1%–100%, and the quantitative recoveries ranged from 91.07% to 111.75%. The results suggested that combination of shotgun proteomics and MRM had potential for the milk species authentication.

B-161 Mass Spectrometric Assay for the Early Diagnosis of Barth Syndrome

Abstract Background Barth syndrome (BTHS) is an inborn error of phospholipid metabolism caused by pathogenic variants in the TAFAZZIN gene locus on a chromosome X. TAFAZZIN encodes a phospholipid transacylase enzyme, tafazzin, that promotes cardiolipin (CL) remodeling. Deficiency of tafazzin activity in BTHS results in accumulation of monolysocardiolipin (MLCL) and decreased abundance of mature CL, making an increased ratio of MLCL to CL as a diagnostic marker for BTHS. 52:2-MLCL to 72:8-CL in dried-blood spot (DBS) has been used for provisional identification BTHS using LC-MS/MS. A development of multiplexed method that targets several species of MLCL and CL can enhance the diagnosis of BTHS. Methods A targeted LC-MS/MS with scheduled multiple reaction monitoring (MRM) mode in negative electrospray ionization was developed using a triple quadrupole MS. DBS from patients with BTHS were obtained at Johns Hopkins University and provided by Barth Syndrome Foundation while control DBS were provided by Biochemical Genetics Laboratory at Mayo Clinic. Internal standards and chloroform:methanol (1:1, v/v) were added to DBS (1/8″ diameter) and sonicated. Supernatants were collected and dried under nitrogen gas. Followed by reconstitution in methanol, samples were subjected to LC-MS/MS analysis. Results More than 20 species of MLCL and CL species having various fatty acyl chains were detected and quantified. Remodeled CL species were decreased while MLCL and nascent CL species were found to be increased in known patients of BTHS (n = 8) compared to controls (n = 8). Notably, several ratios of lipids including 52:2-MLCL/72:8-CL and 54:2-MLCL/70:6-CL showed >300-fold elevation in the patients. Conclusion Various species of MLCL and CL that can be used to diagnose BTHS can be measured from an easily accessible specimen such as DBS using this method.

Simultaneous LC-MS/MS quantification of SGLT2 inhibitors and antipyrine in medium and tissue from human ex vivo placenta perfusions

A liquid chromatography – tandem mass spectrometry (LC-MS/MS) method has been developed to simultaneously measure four sodium glucose co-transporter 2 (SGLT2) inhibitors and the transfer marker antipyrine (ANTI) in perfusion medium and placental tissue collected from ex vivo human placental perfusions. The four SGLT2 inhibitors were empagliflozin (EMPA), dapagliflozin (DAPA), ertugliflozin (ERTU) and canagliflozin (CANA). Chromatographic separation was achieved on an Uptisphere® C18 reversed phase column (50 mm × 4.6 mm × 5 µm) within 2.85 min, using a gradient elution with 10 mM ammonium formate in water (mobile phase A) and acetonitrile (mobile phase B) both with 0.1% formic acid. Analysis of ammonium adduct ions was performed on an AB SCIEX 6500+ triple quadrupole mass spectrometer using positive electrospray ionisation and scheduled multiple reaction monitoring (sMRM). The transitions were m/z 468.00 → 355.20 (EMPA), m/z 426.00 → 167.20 (DAPA), m/z 437.10 → 206.90 (ERTU), m/z 462.00 → 249.00 (CANA) and m/z 189.20 → 55.90 (ANTI).The method was validated according to the European Medicines Agency guidelines and was proven to be selective, linear within a concentration range of 1–1000 µg/L (DAPA, CANA, ANTI) and 1–500 µg/L (EMPA, ERTU), accurate, precise and free of carry-over, instabilities, recovery and matrix effect issues.This newly developed method is suitable to analyse perfusion medium and placenta tissue samples collected during ex vivo human placenta perfusions. It thereby enables quantification of transport across the placental barrier of the SGLT2 inhibitors EMPA, DAPA, ERTU and CANA as well as the transfer marker ANTI.

Comparative Characterization of the Ginsenosides from Six Panax Herbal Extracts and Their In Vitro Rat Gut Microbial Metabolites by Advanced Liquid Chromatography-Mass Spectrometry Approaches.

Ginseng extracts are extensively used as raw materials for food supplements and herbal medicines. This study aimed to characterize ginsenosides obtained from six Panax plant extracts (Panax ginseng, red ginseng, Panax quinquefolius, Panax notoginseng, Panax japonicus, and Panax japonicus var. major) and compared them with their in vitro metabolic profiles mediated by rat intestinal microbiota. Ultrahigh-performance liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) with scheduled multiple reaction monitoring (sMRM) quantitation methods were developed to characterize and compare the ginsenoside composition of the different extracts. After in vitro incubation, 248 ginsenosides/metabolites were identified by UHPLC/IM-QTOF-MS in six biotransformed samples. Deglycosylation was determined to be the main metabolic pathway of ginsenosides, and protopanaxadiol-type and oleanolic acid-type saponins were easier to be easily metabolized. Compared with the ginsenosides in plant extracts, those remaining in six biotransformed samples were considerably fewer after biotransformation for 8 h. However, the compositional differences in four subtypes of the ginsenosides among the six Panax plants became more distinct.

Identification and Quantification of Resveratrol and Its Derivatives in Franconian Wines by Comprehensive Liquid Chromatography–Tandem Mass Spectrometry

Resveratrol and its derivatives are valued compounds in foods such as wine. To date, their occurrence and concentrations in Franconian wines are not known. Thus, the present study investigated resveratrol and its derivatives in Franconian wines. First, comprehensive ultrahigh-performance liquid chromatography (UHPLC)–tandem mass spectrometry methods were applied to identify cis-/trans-resveratrol, cis-/trans-piceid, deoxyrhapontigenin, trans-piceatannol, and cis-/trans-astringin. Second, a sensitive UHPLC–scheduled multiple reaction monitoring method for absolute quantification of the major resveratrol derivatives was developed, validated, and applied to 19 Franconian wine samples. Recovery rates ranged between 95.0 and 109.3% and reproducibility between 2.6 and 10.3%. The limits of detection/quantification were between 0.01 and 0.08 μg/mL. The samples contained 0.07–2.61 μg/mL cis-resveratrol, 0.05–3.82 μg/mL trans-resveratrol, 0.24–9.01 μg/mL cis-piceid, and 0.25–8.30 μg/mL trans-piceid. Additionally, the concentrations of deoxyrhapontigenin (0.02–0.53 μg/mL), trans-piceatannol (0.05–1.35 μg/mL), and trans-astringin (0.04–0.67 μg/mL) were quantified.

Rapid and efficient LC-MS/MS diagnosis of inherited metabolic disorders: a semi-automated workflow for analysis of organic acids, acylglycines, and acylcarnitines in urine.

The analysis of organic acids in urine is an important part of the diagnosis of inherited metabolic disorders (IMDs), for which gas chromatography coupled with mass spectrometry is still predominantly used. Ultra-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for urinary organic acids, acylcarnitines and acylglycines was developed and validated. Sample preparation consists only of dilution and the addition of internal standards. Raw data processing is quick and easy using selective scheduled multiple reaction monitoring mode. A robust standardised value calculation as a data transformation together with advanced automatic visualisation tools are applied for easy evaluation of complex data. The developed method covers 146 biomarkers consisting of organic acids (n=99), acylglycines (n=15) and acylcarnitines (n=32) including all clinically important isomeric compounds present. Linearity with r2>0.98 for 118 analytes, inter-day accuracy between 80 and 120 % and imprecision under 15 % for 120 analytes were achieved. Over 2 years, more than 800 urine samples from children tested for IMDs were analysed. The workflow was evaluated on 93 patient samples and ERNDIM External Quality Assurance samples involving a total of 34 different IMDs. The established LC-MS/MS workflow offers a comprehensive analysis of a wide range of organic acids, acylcarnitines and acylglycines in urine to perform effective, rapid and sensitive semi-automated diagnosis of more than 80 IMDs.

Simultaneous determination of multiple components in rat plasma by UHPLC-sMRM for pharmacokinetic studies after oral administration of Qingjin Yiqi Granules.

As a Traditional Chinese Medicine prescription, Qingjin Yiqi Granules (QJYQ) provides an effective treatment for patients recovering from COVID-19. However, the pharmacokinetics characteristics of the main components of QJYQ in vivo are still unknown. An efficacious ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was developed and validated for the simultaneous determination of 33 components in rat plasma after oral administration of QJYQ. The plasma samples were precipitated with 400µL methanol/acetonitrile (1/1, v/v) and analyzed in scheduled multiple reaction monitoring mode. The linear relationship of the 33 components was good (r > 0.9928). The lower limit of quantification for 33 components ranged from 0.4-60.5ng/mL. The average recoveries and matrix effects of the analytes ranged from 72.9% to 115.0% with RSD of 1.4%-15.0%. All inter-day and intra-day RSDs were within 15.0%. After oral administration (3.15g/kg), the validated approach was effectively applied to the pharmacokinetics of main components of QJYQ. Finally, fifteen main constituents of QJYQ with large plasma exposure were obtained, including baicalin, wogonoside, wogonin, apigenin-7-O-glucuronide, verbenalin, isoferulic acid, hesperidin, liquiritin, harpagide, protocatechuic acid, p-Coumaric acid, ferulic acid, sinapic acid, liquiritin apioside and glycyrrhizic acid. The present research lays a foundation for clarifying the therapeutic material basis of QJYQ and provides a reference for further scientific research and clinical application of QJYQ.

Proteomic Shotgun and Targeted Mass Spectrometric Datasets of Cerebrospinal Fluid (Liquor) Derived from Patients with Vestibular Schwannoma

Vestibular schwannomas are relatively rare intracranial tumors compared to other brain tumors. Data on the molecular features, especially on schwannoma proteome, are scarce. The 41 cerebrospinal fluid (liquor) samples were obtained during the surgical removal of vestibular schwannoma. Obtained peptide samples were analyzed by shotgun LC-MS/MS high-resolution mass spectrometry. The same peptide samples were spiked with 148 stable isotopically labeled peptide standards (SIS) followed by alkaline fractionation and scheduled multiple reaction monitoring (MRM) for quantitative analysis. The natural counterparts of SIS peptides were mapped onto 111 proteins that were Food and Drug Administration (FDA)-approved for diagnostic use. As a result, 525 proteins were identified by shotgun LC-MS/MS with high confidence (at least two peptides per protein, FDR < 1%) in liquor samples. Absolute quantitative concentrations were obtained for 54 FDA-approved proteins detected in at least five experimental samples. Since there is lack of data on the molecular landscape of vestibular schwannoma, the obtained datasets are unique and one of the first in its field.

Quality assessment of Astragali Radix based on pseudo-targeted metabolomics and chemometric approach.

Astragali Radix is widely used because of its dual use in medicine and food, and its quality evaluation is of great importance. In this study, a pseudo-targeted metabolomics approach based on scheduled multiple reaction monitoring was developed, and a total of 114 compounds with good linearity, sensitivity and reproducibility were selected for relative quantification, and the chemical differences between Astragali Radix of different growth patterns were further compared by chemometric analysis. With the help of multivariate and univariate analysis, 26 differential compounds between wild/semi-wild Astragali Radix and cultivated Astragali Radix were determined. Then 5 marker compounds were screened out by lasso regression, and further verified by systematic clustering, random forest, support vector machine, and logistic regression. In addition, malonyl-substituted flavonoids showed relatively higher content in wild/semi-wild Astragali Radix. Thus, the malonyl-substitution was the characteristic for flavonoids in wild/semi-wild Astragali Radix. In conclusion, the application of pseudo-targeted metabolomics and various statistical methods could offer multi-dimensional information for the holistic quality evaluation of Astragali Radix. This article is protected by copyright. All rights reserved.

Development of an enantioselective high-performance liquid chromatography-tandem mass spectrometry method for the quantitative determination of methorphan and its O-demethylated metabolite in human blood and its application to post-mortem samples

In the present work an isocratic enantioselective high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the separation and quantitative determination of dextro - and levo -methorphan and their pharmacologically relevant metabolites, dextrorphan and levorphanol, respectively, in human blood samples. The separation of enantiomers of methorphan and metabolites was performed on the polysaccharide-based chiral column Lux AMP in combination with acetonitrile and 5 mM aqueous ammonium bicarbonate pH 11 in the ratio 50:50 (%, v/v) as mobile phase with the flow rate 1 mL/min. The mass spectrometer was operated in scheduled multiple reaction monitoring (MRM) mode, with four transitions for each dextromethorpan, levomethorphan, dextrorphan and dextromethorphan-d3 and two transitions for each levorphanol, levorphanol-d3 and dextrorphan-d3. Application of this method to human post-mortem blood samples confirmed cases of severe overdosing with dextromethorphan, levomethorphan, and less commonly with both.

A UPLC-MS/MS methodological approach for the analysis of 75 phenethylamines and their derivatives in hair

A rapid ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the targeted analysis of 75 phenethylamines and their derivatives from the hair matrix. The monitored classes of phenethylamines included the 2C series, D series, N-benzyl derivatives, mescaline-derived compounds, MDMA analogs, and benzodifurans. Approximately 20 mg of hair was weighed and pulverized with 0.1% formic acid in methanol by cryogenic grinding. After ultrasonication, centrifugation, and filtration, the supernatant was analyzed by LC–MS/MS operating in the scheduled multiple reaction monitoring mode. Phenethylamines and their derivatives were separated in 13 min on a biphenyl column (2.6 µm, 100 Å, 100 × 3.0 mm) using a gradient eluting mobile phase composed of 0.1% formic acid in water and acetonitrile. The developed and validated method showed good selectivity, sensitivity (LOD: 0.5–10 pg/mg and LOQ: 1–20 pg/mg), linearity (R2 > 0.997), accuracy and precision (< 20%), and stability. The method also showed good recovery and acceptable matrix effects for most of the targeted compounds. This analytical approach was successfully applied for the identification and quantification of phenethylamines in hair from authentic forensic cases.

An efficient method for qualitation and quantitation of multi-components of the herbal medicine Qingjin Yiqi Granules

Qingjin Yiqi Granules (QJYQ) is a Traditional Chinese Medicines (TCMs) prescription for the patients with post-COVID-19 condition. It is essential to carry out the quality evaluation of QJYQ. A comprehensive investigation was conducted by establishing deep-learning assisted mass defect filter (deep-learning MDF) mode for qualitative analysis, ultra-high performance liquid chromatography and scheduled multiple reaction monitoring method (UHPLC-sMRM) for precise quantitation to evaluate the quality of QJYQ. Firstly, a deep-learning MDF was used to classify and characterize the whole phytochemical components of QJYQ based on the mass spectrum (MS) data of ultra-high performance liquid chromatography quadrupole time of flight tandem mass spectrometry (UHPLC-Q-TOF/MS). Secondly, the highly sensitive UHPLC-sMRM data-acquisition method was established to quantify the multi-ingredients of QJYQ. Totally, nine major types of phytochemical compounds in QJYQ were intelligently classified and 163 phytochemicals were initially identified. Furthermore, fifty components were rapidly quantified. The comprehensive evaluation strategy established in this study would provide an effective tool for accurately evaluating the quality of QJYQ as a whole.