High-pressure Mass Spectrometry Research Articles

Exploiting the beneficial effects of Salvia officinalis L. extracts in human health and assessing their activity as potent functional regulators of food microbiota

Salvia officinalis L. has attracted scientific and industrial interest due to its pharmacological properties. However, its detailed phytochemical profile and its correlation with beneficial effects in the human microbiome and oxidative stress remained elusive. To unveil this, S. officinalis was collected from the region of Epirus and its molecular identity was verified with DNA barcoding. Phytochemical profile for both aqueous and ethanol-based extracts was determined by high-pressure liquid chromatography-tandem mass spectrometry and 103 phytochemicals were determined. The effect of S. officinalis extracts as functional regulators of food microbiota by stimulating the growth of Lacticaseibacillus rhamnosus strains and by suppressing evolution of pathogenic bacteria was verified. Furthermore, we recorded that both extracts exhibited a significant cellular protection against H2O2-induced DNA damage. Finally, both extracts exhibited strong inhibitory effect towards LDL oxidation. This study provides a comprehensive characterization of S. officinalis on its phytochemical components as also its potential impact in human microbiome and oxidative stress.

Probing Acid-Induced Compaction of Denatured Proteins by High-Pressure Electrospray Mass Spectrometry.

Further increase in the acidity in the most denaturing acidic solution is known to induce compaction of the fully unfolded protein into a compact molten globule. The phenomenon of "acid-induced folding of proteins" takes place at pH ∼1 in strong acid aqueous solutions with high electrical conductivity and surface tension, a condition that is difficult to handle using conventional electrospray ionization methods for mass spectrometry. Here, high-pressure electrospray ionization (HP-ESI) is used to produce well-resolved mass spectra for proteins in strong acids with pH as low as 1. The compaction of protein conformation is indicated by a large shift in the charge state from high charges to native-like low charges. The addition of salt to the protein in the most denaturing condition also reproduces the compaction effect, thereby supporting the role of anions in this phenomenon. Similar compaction of proteins is also observed in organic solvent/acid mixtures. The charge state of the compacted protein depends on the type of anions that formed ion pairs with a positive charge on the protein. The dissociation of ion pairs during the ionization process forms neutral acids that can be observed by HP-ESI using a soft ion introduction configuration.

Field-portable detection of fentanyl and its analogs: A review.

The need to detect fentanyl and its analogs in the field is an important capability to help prevent unintentional exposure or overdose on these substances, which may result in death. Many portable methods historically used in the field by first responders and other field users to detect and identify other chemical substances, such as hazardous materials, have been applied to the detection and identification of these synthetic opioids. This paper describes field portable spectroscopic methods used for the detection and identification of fentanyl and its analogs. The methods described are automated colorimetric tests including lateral flow assays; vibrational spectroscopy (mid-infrared and Raman); gas chromatography-mass spectrometry; ion mobility spectrometry, and high-pressure mass spectrometry. In each case the background and key details of these technologies are outlined, followed by a discussion of the application of the technology in the field. Attention is paid to the analysis of complex mixtures and limits of detection, including the required spectral databases and algorithms used to interrogate these types of samples. There is also an emphasis on providing actionable information to the (likely) non-scientist operators of these instruments in the field.

Rapid Detection of Volatile Organic Metabolites in Urine by High-Pressure Photoionization Mass Spectrometry for Breast Cancer Screening: A Pilot Study.

Despite surpassing lung cancer as the most frequently diagnosed cancer, female breast cancer (BC) still lacks rapid detection methods for screening that can be implemented on a large scale in practical clinical settings. However, urine is a readily available biofluid obtained non-invasively and contains numerous volatile organic metabolites (VOMs) that offer valuable metabolic information concerning the onset and progression of diseases. In this work, a rapid method for analysis of VOMs in urine by using high-pressure photon ionization time-of-flight mass spectrometry (HPPI-TOFMS) coupled with dynamic purge injection. A simple pretreatment process of urine samples by adding acid and salt was employed for efficient VOM sampling, and the numbers of metabolites increased and the detection sensitivity was improved after the acid (HCl) and salt (NaCl) addition. The established mass spectrometry detection method was applied to analyze a set of training samples collected from a local hospital, including 24 breast cancer patients and 27 healthy controls. Statistical analysis techniques such as principal component analysis, partial least squares discriminant analysis, and the Mann-Whitney U test were used, and nine VOMs were identified as differential metabolites. Finally, acrolein, 2-pentanone, and methyl allyl sulfide were selected to build a metabolite combination model for distinguishing breast cancer patients from the healthy group, and the achieved sensitivity and specificity were 92.6% and 91.7%, respectively, according to the receiver operating characteristic curve analysis. The results demonstrate that this technology has potential to become a rapid screening tool for breast cancer, with significant room for further development.

Pharmacokinetics of Venetoclax Co-Administered with Posaconazole in Patients with Acute Myeloid Leukemia.

The Food and Drug Administration currently approves the combination of hypomethylating agents (HMA), azacytidine or decitabine with venetoclax (VEN) for acute myeloid leukemia (AML) patients aged more than 75 years and for patients unsuitable for intensive chemotherapy. The risk of fungal infection in the early phase of treatment is not negligible; therefore, posaconazole (PCZ) is commonly administered as primary prophylaxis. A drug-drug interaction between VEN and PCZ is well known, but the trend of serum levels of venetoclax when both drugs are overlapped is not clear. In total, 165 plasma samples from 11 elderly AML patients receiving combined treatment with HMA, VEN and PCZ were analyzed by a validated analytical method (high-pressure liquid chromatography-tandem mass spectrometry). Venetoclax trough plasma concentrations were detected during the 3 days of ramp-up as well as on day 7 and day 12 of treatment when the exposure as the area under the plasma concentration-time curve and the accumulation ratio were also calculated. The results were compared with the expected data for 400 mg/dose VEN administered alone-the confirmed high inter-individual variability in pharmacokinetics suggests the need for therapeutic drug monitoring.

Comparison of serum metabolomics in women with breast Cancer Prior to Chemotherapy and at 1 year: cardiometabolic implications

ObjectiveEarly-stage breast cancer (BC) is the second most common malignancy in women, worldwide. Early-detection and treatment advances have led to 5-year survival rates of 90% for early-stage breast cancer. However, the long-term morbidity of breast cancer remains high, with a majority of survivors facing increased risk of cardiometabolic conditions as well as secondary cancers. In particular, African American women with breast cancer experience higher morbidity and mortality than other women. Metabolomics is the comprehensive study of metabolites in biological samples to elucidate the role of monosaccharides, amino acids, and their respective metabolic pathways. Although some studies have found differential metabolites in women with breast cancer compared to normal controls, there has been little study of women with breast cancer across time and the active treatment trajectory. This study examines and compares the serum metabolomic profile of women with BC, prior to initial chemotherapy and at 1 year after inception of chemotherapy.MethodsThis study examined serum metabolites through a secondary analysis of a longitudinal parent study (EPIGEN) of women diagnosed with early-stage BC. Participants were evaluated across 5 time points: prior to their receipt of chemotherapy (T1), at the time of their 4th chemotherapy treatment (T2), 6 months after the initiation of chemotherapy (T3), one year after the initiation of chemotherapy (T4) and two years after the initiation of chemotherapy (T5). This analysis focused on the metabolomic data from 70 participants from T1 to T4. Using ultra high-pressure liquid chromatography high resolution mass spectrometry (UHPLC-HRMS), we performed Friedman Rank Sum Test followed by Nemenyi post-hoc pairwise tests to identify which metabolite levels differed between time points, focusing on metabolites with a Benjamini-Hochberg false discovery rate (FDR) from the overall Friedman test < 0.05 and then specifically examined the p-values from the T1 vs. T4 pairwise comparison.ResultsThe untargeted serum metabolomics yielded a total of 2,395 metabolites identified on the basis of the accurate mass and MS/MS fragmentation, 1,264 of which were significant after Friedman’s test (FDR < 0.05). The analysis then focused on the levels of 124 metabolites from the T1 vs. T4 post-hoc comparison that had a combined FDR < 0.05 and fold change (FC) > 2.0. Metabolite set enrichment analysis (MSEA) as part of Metaboanalyst 3.0 was performed to identify pathways that were significantly altered. The known metabolites identified from the functional analysis were used to evaluate the up and down regulated pathways. The 40metabolites from the Functional Analysis were mainly attributed to amino acids (specifically lysine regulation), fatty acids (particularly unsaturated) and steroid hormone synthesis (lysophosphatidic acid).ConclusionThere were multiple significant changes in the serum metabolomic profile of women with breast cancer at one-year post inception of chemotherapy compared to pre-chemotherapy, most notably associated with lysine degradation, branched-chain amino acid synthesis, linoleic acid metabolism, tyrosine metabolism and biosynthesis of unsaturated fatty acids as the top 5 metabolic pathways. Some of these changes could be associated with metabolic perturbations that are consistent with heightened risk of cardiometabolic morbidity. Our results provide new insights into the mechanisms underlying potential heightened cardiovascular health risks in this population.

Anti-inflammatory effect of a pomegranate extract on LPS-stimulated HepG2 cells

Pomegranate is an important source of bioactive molecules with proven beneficial effects on human health. The aim of this study was to investigate the potential anti-inflammatory effect of a pomegranate extract (PE), obtained from the whole fruit and previously characterized by Reversed Phase-Ultra High-Pressure Liquid Chromatography-High Resolution Mass Spectrometry (RP-UHPLC–HRMS), on HepG2 human hepatocellular carcinoma cells challenged with the lipopolysaccharide (LPS). In LPS-treated cells (1 µg/ml, 24h), the PE treatment (administered at the non-cytotoxic dose of 1 µg/ml, 24h) induced a significant reduction of three key pro-inflammatory cytokines, i.e. interleukin-8 (IL-8), interleukin-1 beta (IL-1β) and tumor necrosis factor-alpha (TNF-α), at both gene expression (as assayed by real-time PCR) and secretion levels (by Enzyme-linked Immunosorbent Assay, ELISA). Although further in vivo studies are needed to prove its efficacy, this preliminary in vitro study suggests that the PE might be useful for ameliorating liver inflammation.

Comprehensive Evaluation of HPLC-MS/MS Method for Quantitation of Seven Anticoagulant Rodenticides and Dicoumarol in Animal Serum.

Anticoagulant rodenticides (ARs) are commonly utilized for controlling rodent populations, however non-target companion and wildlife animals are also exposed. A method was developed for quantitation of seven ARs (chlorophacinone, coumachlor, bromadiolone, brodifacoum, difethialone, diphacinone, and warfarin) and dicoumarol (a naturally occurring anticoagulant) in animal serum. Analytes were extracted with 10% (v/v) acetone in methanol and analyzed by reverse phase high-pressure liquid chromatograph-tandem mass spectrometry (HPLC-MS/MS) using electrospray ionization (negative mode) combined with multiple reaction monitoring (MRM). In-house method validation in the originating laboratory using non-blinded samples revealed method limits of quantitation at 2.5 ng/mL for all analytes. Inter-assay accuracy ranged 99-104% and relative standard deviation ranged 3.5-20.5%. Method performance was then verified in the originating laboratory during an exercise organized by an independent party using blinded samples. The method was successfully transferred to two naïve laboratories and further evaluated for reproducibility among three laboratories by means of Horwitz ratio (HorRat(R)) values. Such extensive validation provides a high degree of confidence that the method is rugged, robust, and will perform as expected if used by others in the future.

25-Hydroxyvitamin D: A potential marker of the incidence of osteoporosis and sarcopenia in diabetic mellitus patients

Purpose: To investigate the feasibility of using the 25-hydroxyvitamin D (25(OH)D) levels in assessing osteoporosis (OP) and sarcopenia in elderly patients with diabetes mellitus (DM).&#x0D; Methods: One hundred and forty elderly patients with DM were allocated to DM (n = 40), OP (n = 50), and DM + OP (n = 50) groups. Their clinical data were collected, and muscle function assessed. The levels of fasting insulin, glycated hemoglobin (HbAlc), t-P1NP, and 25(OH)D were determined by chemiluminescent microparticle immunoassay, high-pressure liquid chromatography, electrochemical immunoassay, and liquid chromatography-tandem mass spectrometry, respectively.&#x0D; Results: The 25(OH)D levels were lowest in DM + OP group, followed by the OP group, and highest in the DM group (p &lt; 0.05). The DM + OP group showed significantly lower gait speed and grip strength than the OP and DM groups (p &lt; 0.05). The diagnostic AUC of 25(OH)D for OP and sarcopenia were 0.9733 (95 % CI: 0.9274 - 1.000, p &lt; 0.001) and 0.9866 (95 % CI: 0.9632 - 1.000, p &lt; 0.001). Patients with sarcopenia had significantly lower 25(OH)D and t-PINP levels than patients without sarcopenia (p &lt; 0.05). Logistic regression analysis revealed that the 25(OH)D level, duration of DM, and age were independent risk factors for sarcopenia (p &lt; 0.05).&#x0D; Conclusion: Patients with DM and OP exhibit increase in the incidence of sarcopenia. Serum 25(OH)D may be used as a potential marker for the incidence of OP and sarcopenia in DM patients.

Effects of d-ribose on human erythrocytes: Non-enzymatic glycation of hemoglobin, eryptosis, oxidative stress and energy metabolism

d-Ribose is not only an important component of some biomacromolecules, but also an active pentose with strong reducibility and non-enzymatic glycation ability. Previous studies reported the diverse role of d-ribose in different cells. In this study, the effects of d-ribose on non-enzymatic glycation of hemoglobin (Hb), as well as eryptosis, oxidative stress and energy metabolism of erythrocytes were observed by molecular fluorescence spectrophotometry, multi-wavelength spectrophotometry, high-pressure liquid chromatography (HPLC), mass spectrometry (MS) and flow cytometer. The results showed that d-ribose had the strongest non-enzymatic glycation ability to Hb in vitro when compared with other monosaccharides, and could enter the erythrocytes in a concentration-dependent manner, which was not inhibited by the specific glucose transporter 1 (GLUT1) inhibitor WZB117. In addition, d-ribose incubation increased the HbA1c, hemolysis, eryptosis, and ROS level of erythrocytes significantly more than that of d-glucose, however, no changes were observed in the levels of ATP, NADPH, and other intermediate energy metabolites in d-ribose treatment. Therefore, the strong non-enzymatic glycation ability of d-ribose may play an important role in erythrocyte damage.

Synthesis and Characterization of a Novel Peptide Targeting Human Tenon Fibroblast Cells To Modulate Fibrosis: An Integrated Empirical Approach.

Fibrosis is the primary factor influencing the prognosis of glaucoma post-trabeculectomy surgery, an eye condition characterized by increased intraocular pressure (IOP). Despite advancements in surgical procedures and aftercare, it continues to be a serious impediment. During the clinical intervention of scarring, fibrosis is managed by using topical application of combined antifibrotic drugs (mitomycin C). But still, scarring remains a key problem due to minimal drug penetration and nonbioavailability. In this study, we synthesized a cell-specific peptide for modulating scarring in human tenon fibroblasts undergoing epithelial-mesenchymal transition (EMT). The peptide was also conjugated with mitomycin C in order to investigate the effect of the drug conjugation on human tenon fibroblasts from the nanofiber composite system and to evaluate the fibrosis process. Peptide VRF2019 was identified using a subtractive proteomics approach, including solubility, cell penetration, and amphipathic properties. The peptide structure was determined using circular dichroism spectroscopy. The peptide and drug was conjugated using N-ethyl-N'-(3-(dimethylamino)propyl) carbodiimide/N-hydroxysuccinimide (EDC-NHS) chemistry, and the conjugation efficiency was evaluated using high-pressure liquid chromatography. The conjugated product and polycaprolactone (PCL) were electrospun to form a composite nanofiber, which was tested for cytotoxicity and drug release on human tenon fibroblast cells. The modeled VRF2019 peptide structure formed an α-helical structure with all residues spanning the allowed regions of the Ramachandran plot. Subsequent molecular dynamics simulations also demonstrated its membrane penetration potential. The peptide uptake was also studied in human tenon fibroblast cells. High-pressure liquid chromatography (HPLC) and mass spectrometry measurements confirmed peptide-drug conjugation and stability. Furthermore, scanning electron microscopy (SEM) investigation revealed the structure and size of the PCL composite nanofiber. We infer from early research that the PCL composite nanofiber matrix can greatly increase drug delivery and bioavailability.

Spectrometers in Wonderland: Shrinking, Shrinking, Shrinking

This article gives a brief overview of the major portable techniques: those based on optical spectroscopy techniques, including near-infrared (NIR), mid-infrared (mid-IR), and Raman spectroscopy; mass spectrometry (MS) systems, including high-pressure MS (HPMS), gas chromatography–MS (GC–MS), ion mobility spectrometry (IMS); elemental techniques, such as X-ray fluorescence (XRF) and laser-induced breakdown spectroscopy (LIBS); and emerging miniaturized techniques like nuclear magnetic resonance (NMR). The above are all “conventional” spectroscopic techniques and reduced to a rugged portable format, containing self-contained data systems. They provide specific and actionable information to their operators working with them outside the laboratory—in the field—and these instruments have well-defined value propositions. A recent development is the availability of low cost (&lt;$100) multispectral sensors operating in the visible and NIR regions. This low cost enables the sensors to be embedded into consumer products, such as smart “white goods” appliances, personal care, fitness products, and even “wearables” products. In the future, miniature and portable spectrometers will be ubiquitous—outside the laboratory, and in your home and pocket.

Creatine Deficiency Syndromes: Comparison of Screening Methods and Characterization of Four Novel Intronic Variants

BackgroundCerebral creatine deficiency syndromes (CCDS) are disorders affecting creatine synthesis or transport. Several methods have been developed to measure creatine and guanidinoacetate (GAA) in different body fluids including methods based on gas chromatography-mass spectrometry (GC–MS) and High-pressure liquid chromatography mass spectrometry (HPLC-MS). The diagnosis of CCDS is then confirmed by sequencing of creatine biosynthesis genes guanidinoacetate methyltransferase (GAMT) and Arginine: glycine amidinotransferase (GATM) and creatine transporter gene solute carrier family 6 member 8 (SLC6A8) or by functional enzymatic assay. The aim of the current study was to find the most reliable and accurate screening method for CCDS by comparing methods using Nuclear Magnetic Resonance spectroscopy (NMR), GC–MS and HPLC-MS. Additionally, this study was performed to estimate the prevalence of CCDS in a cohort of Egyptian patients and potentially to discover novel variants. Subjects and methodsThe study was conducted on 150 subjects with clinical signs and symptoms consistent with CCDS. Metabolic profiling of urine samples was performed using three techniques: 1) GC–MS 2) Ultra high-pressure (or performance) liquid chromatography – Tandem Mass Spectrometry (UHPLC- MS/MS) and 3) NMR. ResultsThe linearity of peak areas for creatine and GAA by UHPLC-MS/MS and NMR covered and exceeded the ranges normally found in urine. The limit of quantification and the inter-day precision results for creatine and GAA were more robust by UHPLC-MS/MS than NMR. Ten cases were identified as being positive for CCDS by our analytical approaches and underwent next generation sequencing (NGS) for GAMT, GATM and SLC6A8 genes. NGS was performed and confirmed one patient with one likely Pathogenic variant in GAMT gene: (NC_000019.10:g.1401317C > G, NP_000147.1:p.Ala54Pro). Additionally, we describe four novel intronic variants in the GATM gene: c.1043-357del and c.1043-357_1043-356insT, and were predicted to activate cryptic acceptor site with potential alteration of splicing, c.979-227G > A was found to significantly alter the Exon Splice Enhancer (ESE) xon Splice Silencer (ESS) motifs ratio and c.1042 + 262del which was found to have no implications on splicing. ConclusionsBoth UHPLC-MS/MS and NMR spectroscopy are comparable to GC–MS in screening for CCDS. Nonetheless, the UHPLC-MS/MS method had better performance than NMR spectroscopy. Additionally, Sequencing of the full length of GATM, GAMT, and SLC6A8 genes is needed to identify intronic variants that could cause CCDS via affecting splice sites.

Negative Ion Photoelectron Spectra of Deprotonated Benzonitrile Isomers via Computation of Franck-Condon Factors.

Negative ion photoelectron spectra of ortho (o-), meta (m-), and para (p-) deprotonated benzonitrile (o-, m-, p-C6H4(CN)-) isomers as well as the associated thermochemical values corresponding to deprotonation at o-, m-, and p-positions in C6H5(CN) are presented. Quantum mechanical results based on the density functional theory (DFT) utilizing the aug-cc-pVQZ basis set indicate that the o-, m-, p-C6H4(CN)● radicals have electron affinity values (EAs) of 1.901, 1.778, and 1.789 eV, respectively. The computed Franck-Condon (FC) factors give rise to o-, m-, and p-C6H4(CN)- negative ion spectra with FC active ring distortion vibrational modes with harmonic vibrational frequencies of ∼450, 760, and 1000 cm-1 as the dominant vibrational progressions. Deprotonation at the o-, m-, and p-positions in C6H5(CN) results in calculated gas-phase acidity values (ΔacidH298Ko) of 383.9, 385.7, and 385.3 kcal mol-1, respectively. The calculated ΔacidH298Ko is in close agreement with the previously reported high-pressure mass spectrometry experimental value of 383.4.0 ± 4.4 kcal mol-1. The computed ΔacidH298Ko and EAs are utilized to estimate the bond dissociation energy (DH298(H-C6H4CN)) associated with the formation o-, m-, and p-C6H4(CN)● using the negative ion thermochemical cycle: DH298(C6H5CN) = ΔacidH298Ko (H-C6H4(CN) + EA (C6H5CN)● - IP(H). The respective values of DH298(H-C6H4CN) corresponding to the formation of ortho, meta, and para C6H4(CN) radicals are 114.15, 113.11, and 113.51 kcal mol-1.

Abstract 529: SNAQ-SEQ™: Use of synthetic internal standards in conjunction with poisson exact test to call variants in contrived circulating tumor DNA specimens

Abstract Background: Targeted next generation sequencing (NGS) analysis of circulating tumor (ct) DNA promises to significantly advance targeted therapy and potentially early diagnosis of cancer. However, targeted NGS has poor accuracy for calling known variants with VAF below 0.5%. The goal of this study from the FDA-Sequencing and Quality Control Phase 2 (SEQC2) consortium was to develop and validate bioinformatic and biostatistical methods that enable incorporation of synthetic competitive internal standards (IS) in targeted NGS analysis of actionable tumor mutations. Methods: A synthetic IS spike-in was designed for each actionable mutation target, suitable for use in NGS following targeted PCR or hybrid-capture enrichment and either with unique molecular index (UMI) or non-UMI library preparation. Contrived ctDNA reference samples developed by the SEQC2 consortium containing actionable mutations at known variant allele fraction were used. An aliquot of each sample was mixed with a mixture of IS. Following Illumina TST170 enrichment and library preparation, each library was sequenced, then native template (NT) sequences were separated from IS sequences bioinformatically. In SNAQ-SEQ™ analysis, Poisson Exact Test (PET) analysis was used to calculate the statistical difference between each sample library NT variant VAF and respective IS variant VAF. Analysis was based on NT variant count and position coverage (i.e., copies recovered in library preparation) and IS count and position coverage. PET performed an exact test of a simple null hypothesis about the ratio between two rate parameters in Poisson distribution. Results: Stochastic sampling effect on IS error detection was minimized by ensuring an IS/NT ratio of 2.5 or greater. Under the specified conditions, in which a minimum of two NT variant observations were required for a call, the IS was able to estimate NGS background error for each sample when a minimum IS:NT ratio of 2.5:1 was used. Without use of IS information, the Illumina pipeline called 73% (41/56) of known TP variants in the 0.1% - 0.3% VAF range. In contrast, SNAQ-SEQ™ analysis (PET analysis of IS and NT information) increased TP detection sensitivity to 86% (48/56), for a 13% increase in sensitivity, with no false positives. Conclusion: Following mixture of contrived ctDNA reference samples with IS, PET analysis enabled calculation of technical error rate, limit of blank, and limit of detection for each variant at each nucleotide position, in each sample. Using this SNAQ-SEQ™ analysis, true positive mutations with variant allele fraction too low for detection by current practice were detected with this method, thereby increasing sensitivity. SNAQ-SEQ™ provides QC that is already standard operating procedure in clinical laboratories for analysis by high pressure liquid chromatography and mass spectrometry. Citation Format: James C. Willey, Erin Crawford, Daniel J. Craig, Joshua Xu, Nathan Haseley, Jennifer Lococo, Tom Morrison. SNAQ-SEQ™: Use of synthetic internal standards in conjunction with poisson exact test to call variants in contrived circulating tumor DNA specimens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 529.

Bayesian model calibration for vacuum-ultraviolet photoionisation mass spectrometry

We investigate the calibration of an instrument model for a high-pressure chemical reactor and vacuum ultraviolet photoionisation mass spectrometry apparatus, using a collection of ‘static’ mass spectrometry data (i.e. from unreactive samples). A Bayesian calibration method is applied to characterise the uncertainty of the model parameters, the associated model error, and the predicted mass spectrum. A partitioning algorithm is introduced to decompose the mass spectrum into several independent calibration problems, making the analysis of complex mass spectrometry data sets tractable. The a priori mass spectrometer model poorly predicts some portions of the ion time-of-flight spectrum due to both, parametric (i.e. uncertain parameter values) and mechanistic (i.e. missing physics) inadequacies; however, by incorporating an additive model error the prediction model is vastly improved and closely replicates the measured spectrum. We found that the model calibration and additive model error reveals a set of experimental problems, including chemical contamination of the sample, the presence of high harmonics of the ionising radiation, and a systematic fluctuation in ion flight times due to instability of the ion optics power supplies. This approach results in a calibrated instrument model with well-defined parameter uncertainties, which is critical to the analysis of ‘dynamic’ data (i.e. from reacting gas samples).

Age-dependent reference intervals in children for the disialoganglioside, GD2, a circulating tumor biomarker for the childhood cancer, neuroblastoma

BackgroundThe disialoganglioside GD2 is a circulating tumor biomarker for the childhood cancer, neuroblastoma. This study establishes reference intervals for GD2 concentration in children within the age range where neuroblastoma commonly occurs. MethodsLeftover plasma samples taken for routine clinical laboratory tests from children without cancer were collected and assayed for the 18-carbon fatty acid chain length lipoform of GD2 using a validated high-pressure liquid chromatography tandem mass spectrometry method with a lower limit of quantification of 3 nM. Samples were stratified into 5 age cohorts (0–6 months, 6–12 months, 12–36 months, 3–10 years and > 10 years). Non-parametric statistical methods were used to define the upper bound of the reference interval for each age cohort. ResultsGD2 was measurable in 90% of samples from children < 10 years of age and GD2 concentration was age-dependent, peaking at 9 months followed by a gradual decline. GD2 was below the lower limit of quantification in 55% of samples in the > 10 years cohort. Upper bounds of reference intervals were 15.5 nM in 0–6 month cohort, 35.1 nM in 6–12 month cohort, 24.9 nM in 12–36 month cohort, 18.4 in 3–10 year cohort and 10.4 nM in > 10 year cohort. ConclusionsAge-dependent reference intervals were defined for circulating GD2 in children. GD2 concentration was highest in the 6–12 month age cohort, which is below the age of most children with high-risk neuroblastoma. The peak GD2 concentration at 9 months may reflect neurodevelopmental events in the brain.

Pharmacokinetics of Synthetic Cathinones Found in Bath Salts in Mouse Brain and Plasma Using High-Pressure Liquid Chromatography-Tandem Mass Spectrometry.

Approximately 10 years ago, "bath salts" became popular as legal alternatives to the psychostimulants cocaine and the amphetamines. These products contained synthetic cathinones, including 3,4-methylenedioxypyrovalerone (MDPV), 4-methylmethcathinone (mephedrone), and 3,4-methylenedioxymethcathinone (methylone). Most preclinical investigations have only assessed the effects of these synthetic cathinones independently; however, case reports and Drug Enforcement Administration (DEA) studies indicate that bath salts contain mixtures of these substances. In this study, we examine the pharmacokinetic interactions of the drug combination. We hypothesized that combined exposure to MDPV, mephedrone, and methylone would result in increased drug concentrations and enhanced total drug concentrations when compared to individual administration. Adolescent male Swiss-Webster mice were injected intraperitoneally with either 10 mg/kg MDPV, 10 mg/kg mephedrone, 10 mg/kg methylone, or 10 mg/kg combined MDPV, mephedrone, and methylone. Following injection, brains and plasma were collected at 1, 10, 15, 30, 60, and 120 min. Drugs were extracted via solid-phase extraction, and concentrations were determined using a previously published high-pressure liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method. All drugs crossed the blood-brain barrier quickly. For methylone, the maximal concentration (Cmax) and the total drug exposure [as represented by the area under the concentration-time curve (AUC)] were significantly higher when combined with mephedrone and MDPV in both matrices (2.89-fold increase for both Cmax and AUC with combined treatment). For mephedrone, the Cmax was unchanged, but the AUC in brain was increased when in combination by approximately 34%. Interestingly, for MDPV, the Cmax was unchanged, yet the AUC was higher when MDPV was administered individually (there was a 62% decrease in AUC with combined treatment). The pharmacokinetics of methylone, mepedrone, and MDPV are altered when the drugs are used in combination. These data provide insight into the consequences of co-exposure to synthetic cathinones in popular bath salt products.

Inhibition Kinetics and Mechanism of Glutathione and Quercetin on Acrylamide in the Low-Moisture Maillard Systems

The inhibition kinetics of glutathione (GSH) and quercetin on acrylamide (AA) formation in the low-moisture Maillard systems were investigated at 180°C. The inhibition rates in an equal-molar asparagine-glucose (Asn-Glc) system were higher than those in an asparagine-fructose (Asn-Fru) system, and the maximum inhibition rates for AA were 57.75% with 10-2 mol L-1 GSH and 51.38% with 10-1 mol L-1 quercetin. The Logistic-Index dynamic model and two consecutive simplified first-order kinetic models were well fitted to the changes of AA in the Asn-Glc system. The kinetics results suggested that the predominant inhibition effect of GSH on AA could be attributed to the competitive reaction between GSH and Asn for the consumption of Glc. The kinetic results and high-pressure liquid chromatography-tandem mass spectrometry analysis of the inhibitory effect of quercetin on AA indicated that quercetin might mitigate AA through the binding reaction of quercetin decomposition products and Maillard intermediate products. These experimental results provide theoretical data that may be useful to control the formation of AA during food thermal processing.

Pharmacokinetics of a cytosine arabinoside subcutaneous protocol in dogs with meningoencephalomyelitis of unknown aetiology.

Cytosine arabinoside (CA) is a commonly used treatment for dogs with meningoencephalomyelitis of unknown aetiology (MUE) with various proposed protocols, many requiring 24hours (h) of hospitalization or two visits within 24h. This is a unidirectional study evaluating the pharmacokinetics of a CA subcutaneous (SC) protocol and a standard constant rate infusion (CRI) protocol in 8 dogs with MUE. Dogs received the CRI (200mg/m2 IV over 24h), followed by a SC protocol (50mg/m2 every 2h for 4 treatments) four weeks later. Plasma CA concentrations were measured by high-pressure liquid chromatography-tandem mass spectrometry (HPLC-MS). Median peak CA concentration for the SC protocol (3.40µg/ml, range 1.60-9.70µg/ml) was significantly higher than the CRI (1.09µg/ml, range 0.77-1.67µg/ml; p=.02). Median concentration at 1h and 8h following initiation of treatment was significantly higher for the SC protocol (CA1 2.28µg/ml, range 0.97-2.67; CA8 1.83µg/ml, range 0.77-2.84) compared to the CRI (CA1 0.01µg/ml, range 0-0.45; CA8 0.74µg/ml, range 0.67-1.11; p=.01). While the PK properties of CA when administered as a CRI has been previously investigated, this study demonstrated that CA when administered via repeated 50mg/m2 injections every 2h over an 8-h period, provided sustained plasma levels above its therapeutic target and for a significantly longer duration of time than did a standard CRI protocol.