Gastric Fluid Metabolomics Predicting the Need for Surfactant Replacement Therapy in Very Preterm Infants Results of a Case-Control Study.

Respiratory distress syndrome (RDS) is a major morbidity of prematurity. In this case-control study, we prospectively evaluated whether untargeted metabolomic analysis (gas chromatography-mass spectrometry) of the gastric fluid could predict the need for surfactant in very preterm neonates. 43 infants with RDS necessitating surfactant (cases) were compared with 30 infants who were not treated with surfactant (controls). Perinatal-neonatal characteristics were recorded. Significant differences in gastric fluid metabolites (L-proline, L-glycine, L-threonine, acetyl-L-serine) were observed between groups, but none could solely predict surfactant administration with high accuracy. Univariate analysis revealed significant predictors of surfactant administration involving gastric fluid metabolites (L-glycine, acetyl-L-serine) and clinical parameters (gestational age, Apgar scores, intubation in the delivery room). Multivariable models were constructed for significant clinical variables as well as for the combination of clinical variables and gastric fluid metabolites. The AUC value of the first model was 0.69 (95% CI 0.57-0.81) and of the second, 0.76 (95% CI 0.64-0.86), in which acetyl-L-serine and intubation in the delivery room were found to be significant predictors of surfactant therapy. This investigation adds to the current knowledge of biomarkers in preterm neonates with RDS, but further research is required to assess the predictive value of gastric fluid metabolomics in this field.

An interview with Bioanalysis: speaking with the 2023 Reid bursary award winners - part 2.

The 25th edition of the International Reid Bioanalytical Forum (REID) was held at the Cambridge Belfry (Cambourne, UK) between 4 and 7 September 2023 and hosted approximately 100 delegates, the majority of whom were attending the event for their first time. REID encourages early-career researchers to present their work and have a bursary program to help provide them support. At the 2023 event, REID welcomed 15 bursary winners to provide them with the opportunity to participate in their first international meeting, network with their peers and make their first oral, or poster presentation. The bursary winners also had the opportunity to interview with the Bioanalysis journal and their responses to the interview questions are transcribed below in this second part of two.

Detection of 26 Drugs of Abuse and Metabolites in Quantitative Dried Blood Spots by Liquid Chromatography-Mass Spectrometry.

A method was developed for the determination of 26 drugs of abuse from different classes, including illicit drugs in quantitative dried blood spots (qDBSs), with the aim to provide a convenient method for drug testing by using only 10 µL of capillary blood. A satisfactory limit of quantification (LOQ) of 2.5 ng/mL for 9 of the compounds and 5 ng/mL for 17 of the compounds and a limit of detection (LOD) of 0.75 ng/mL for 9 of the compounds and 1.5 ng/mL for 17 of the compounds were achieved for all analytes. Reversed-phase liquid chromatography was applied on a C18 column coupled to MS, providing selective detections with both +ESI and -ESI modes. Extraction from the qDBS was performed using AcN-MeOH, 1:1 (v/v), with recovery ranging from 84.6% to 106%, while no significant effect of the hematocrit was observed. The studied drugs of abuse were found to be stable over five days under three different storage conditions (at ambient temperature 21 °C, at -20 °C, and at 35 °C), thus offering a highly attractive approach for drug screening by minimally invasive sampling for individuals that could find application in forensic toxicology analysis.

Dried urine spot (DUS) applied for sampling prior to the accurate HILIC-MS/MS determination of 14 amino acids.

Urine amino acid analysis has proven valuable for an array of clinical or nutritional studies. However, transportation of liquid urine sample shows certain disadvantages, such as possible leakage, need for cold chain and thus higher costs for their transport. Utilization of dried urine spots (DUS) can offer an interesting alternative. In the present study, a method was developed for the determination of 14 amino acids in DUS including the testing of in-house collection device and drying of the sample before analysis. Normal filter paper was tested as the means for sample collection. Absorption and extraction experiments were performed on 3 different types of filter paper, with 3 different extraction solvents and two different solvent volumes. The solvents used were mixtures of common analytical solvents (methanol, water, acetonitrile) using total volumes of 1 mL and 1.5 mL. Finally, 1 mL of acetonitrile: methanol: water 40:40:20 (v/v/v) was chosen as the optimal system. Analysis was performed on a UHPLC-MS system, using stable isotope labeled internal standards. Method validation included the study of limits of detection (LOD) and quantification (LOQ), linearity ranges, precision, matrix effect, extraction recovery, precision, and stability for each analyte. The obtained results were satisfactory, thus enabling application of the proposed method as an alternative to the analysis of liquid urine. Further utilization of DUS can offer advantages by enabling patient centric sampling even in long distances far from the analytical laboratories.

The Contribution of Lipidomics in Ovarian Cancer Management: A Systematic Review.

Lipidomics is a comprehensive study of all lipid components in living cells, serum, plasma, or tissues, with the aim of discovering diagnostic, prognostic, and predictive biomarkers for diseases such as malignant tumors. This systematic review evaluates studies, applying lipidomics to the diagnosis, prognosis, prediction, and differentiation of malignant and benign ovarian tumors. A literature search was performed in PubMed, Science Direct, and SciFinder. Only publications written in English after 2012 were included. Relevant citations were identified from the reference lists of primary included studies and were also included in our list. All studies included referred to the application of lipidomics in serum/plasma samples from human cases of OC, some of which also included tumor tissue samples. In some of the included studies, metabolome analysis was also performed, in which other metabolites were identified in addition to lipids. Qualitative data were assessed, and the risk of bias was determined using the ROBINS-I tool. A total of twenty-nine studies were included, fifteen of which applied non-targeted lipidomics, seven applied targeted lipidomics, and seven were reviews relevant to our objectives. Most studies focused on the potential application of lipidomics in the diagnosis of OC and showed that phospholipids and sphingolipids change most significantly during disease development. In conclusion, this systematic review highlights the potential contribution of lipids as biomarkers in OC management.

A Cohort Study of Gastric Fluid and Urine Metabolomics for the Prediction of Survival in Severe Prematurity.

Predicting survival in very preterm infants is critical in clinical medicine and parent counseling. In this prospective cohort study involving 96 very preterm infants, we evaluated whether the metabolomic analysis of gastric fluid and urine samples obtained shortly after birth could predict survival in the first 3 and 15 days of life (DOL), as well as overall survival up to hospital discharge. Gas chromatography-mass spectrometry (GC-MS) profiling was used. Uni- and multivariate statistical analyses were conducted to evaluate significant metabolites and their prognostic value. Differences in several metabolites were identified between survivors and non-survivors at the time points of the study. Binary logistic regression showed that certain metabolites in gastric fluid, including arabitol, and succinic, erythronic and threonic acids, were associated with 15 DOL and overall survival. Gastric glyceric acid was also associated with 15 DOL survival. Urine glyceric acid could predict survival in the first 3 DOL and overall survival. In conclusion, non-surviving preterm infants exhibited a different metabolic profile compared with survivors, demonstrating significant discrimination with the use of GC-MS-based gastric fluid and urine analyses. The results of this study support the usefulness of metabolomics in developing survival biomarkers in very preterm infants.

Optimization of Carob Products Preparation for Targeted LC-MS/MS Metabolomics Analysis.

Carob (Ceratonia siliqua) is an exceptional source of significant bioactive compounds with great economic importance in the Mediterranean region, where it is widely cultivated. Carob fruit is used for the production of a variety of products and commodities such as powder, syrup, coffee, flour, cakes, and beverages. There is growing evidence of the beneficial effects of carob and the products made from it on a range of health problems. Therefore, metabolomics could be used to explore the nutrient-rich compounds of carob. Sample preparation is a crucial step in metabolomics-based analysis and has a great impact on the quality of the data obtained. Herein, sample preparation of carob syrup and powder was optimized, to enable highly efficient metabolomics-based HILIC-MS/MS analysis. Pooled powder and syrup samples were extracted under different conditions by adjusting pH, solvent type, and sample weight to solvent volume ratio (Wc/Vs). The metabolomics profiles obtained were evaluated using the established criteria of total area and number of maxima. It was observed that the Wc/Vs ratio of 1:2 resulted in the highest number of metabolites, regardless of solvent type or pH. Aqueous acetonitrile with a Wc/Vs ratio of 1:2 satisfied all established criteria for both carob syrup and powder samples. However, when the pH was adjusted, basic aqueous propanol 1:2 Wc/Vs and acidic aqueous acetonitrile 1:2 Wc/Vs provided the best results for syrup and powder, respectively. We strongly believe that the current study could support the standardization of the metabolomics sample preparation process to enable more efficient LC-MS/MS carob analysis.

Ceramides biomarkers determination in quantitative dried blood spots by UHPLC-MS/MS.

A method was developed for the analysis of four ceramide species; namely C16:0, C18:0, C24:0 and C24:1 in quantitative Dried Blood Samples (qDBS) by LC-MS/MS and validated with the aim to give prominence to an interesting application of at-home blood microsampling for health monitoring. Ceramides, being key-role metabolites implicated in regulation of diverse cellular processes have been considered as emerging biomarkers for different disease states, such as cardiovascular diseases, type 2 diabetes and others. Here, Capitainer device was utilized to provide accurate and consistent volumes of samples, ideal for accurate determinations. The method requires a 10 µL sample offering duplicate analysis by device, is quick and enables the sample collection by distance as it was proved that ceramides under study were stable at various conditions, including RT. Intra and inter-day accuracy of the determination were estimated between 87.6% - 113% and 90.6% -113%, respectively, while intra- and inter-day precision were calculated from 0.2% to 9.9% %RSD and 0.1% - 8.0% %RSD, respectively. The data acquired by ten healthy individuals indicated that circulating ceramides are at higher levels in whole blood taken from the fingertip in comparison to the reported values in plasma or serum.

Tigecycline Pharmacokinetic and Pharmacodynamic Profile in Patients with Chronic Obstructive Pulmonary Disease Exacerbation.

BACKGROUND: We aimed to evaluate the pharmacokinetic profile of tigecycline in plasma and its penetration to sputum in moderately ill patients with an infectious acute exacerbation of chronic obstructive pulmonary disease (COPD). METHODS: Eleven patients hospitalized with acute respiratory failure due to an acute COPD exacerbation with clinical evidence of an infectious cause received tigecycline 50 mg twice daily after an initial loading dose of 100 mg. Blood and sputum samples were collected at steady state after dose seven. RESULTS: In plasma, mean Cmax pl was 975.95 ± 490.36 ng/mL and mean Cmin pl was 214.48 ±140.62 ng/mL. In sputum, mean Cmax sp was 641.91 ± 253.07 ng/mL and mean Cmin sp was 308.06 ± 61.7 ng/mL. In plasma, mean AUC 0-12 pl was 3765.89 ± 1862.23 ng*h/mL, while in sputum mean AUC 0-12 sp was 4023.27 ± 793.37 ng*h/mL. The mean penetration ratio for the 10/11 patients was 1.65 ± 1.35. The mean Free AUC0-24 pl/MIC ratio for Streptococcus pneumoniae and Haemophilus influenzae was 25.10 ± 12.42 and 6.02 ± 2.97, respectively. CONCLUSIONS: Our findings support the clinical effectiveness of tigecycline against commonly causative bacteria in COPD exacerbations and highlight its sufficient lung penetration in pulmonary infections of moderate severity.

Untargeted Metabolomics Pilot Study Using UHPLC-qTOF MS Profile in Sows' Urine Reveals Metabolites of Bladder Inflammation.

Urinary tract infections (UTI) of sows (characterized by ascending infections of the urinary bladder (cyst), ureters, and renal pelvis), are major health issues with a significant economic impact to the swine industry. The current detection of UTI incidents lacks sensitivity; thus, UTIs remain largely under-diagnosed. The value of metabolomics in unraveling the mechanisms of sow UTI has not yet been established. This study aims to investigate the urine metabolome of sows for UTI biomarkers. Urine samples were collected from 58 culled sows from a farrow-to-finish herd in Greece. Urine metabolomic profiles in 31 healthy controls and in 27 inflammatory ones were evaluated. UHPLC-qTOF MS/MS was applied for the analysis with a combination of multivariate and univariate statistical analysis. Eighteen potential markers were found. The changes in several urine metabolites classes (nucleosides, indoles, isoflavones, and dipeptides), as well as amino-acids allowed for an adequate discrimination between the study groups. Identified metabolites were involved in purine metabolism; phenylalanine; tyrosine and tryptophan biosynthesis; and phenylalanine metabolism. Through ROC analysis it was shown that the 18 identified metabolite biomarkers exhibited good predictive accuracy. In summary, our study provided new information on the potential targets for predicting early and accurate diagnosis of UTI. Further, this information also sheds light on how it could be applied in live animals.

Plasma Ceramide Concentrations in Full-Term Pregnancies Complicated with Gestational Diabetes Mellitus: A Case-Control Study.

Ceramides, a sphingolipid group that acts as a messenger in cellular differentiation, proliferation, apoptosis and senescence, have been associated with cardiovascular disease and type 2 diabetes. The evidence for an association between ceramides and gestational diabetes mellitus (GDM) is scarce. This case-control study aimed to compare women with GDM with healthy, pregnant women in terms of plasma ceramide concentrations at the time of delivery. Ninety-two pregnant women were included in this case-control study, 29 in the GDM group and 63 in the control group. All women were admitted to a tertiary academic hospital for a full-term delivery. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was applied for the quantification of four molecular ceramides, namely Cer d18:1/16:0 (Cer16:0), Cer d18:1/18:0 (Cer18:0), Cer d18:1/24:0 (Cer24:0) and Cer d18:1/24:1 (Cer24:1) in plasma samples. The raw chromatographic data obtained from the LC-MS/MS analysis were processed using Analyst SCIEX (AB Sciex Pte. Ltd., USA). In a univariate statistical analysis, Cer24:0 concentration was significantly lower in the GDM group compared with the control group (p = 0.01). The present study demonstrated lower Cer24:0 concentrations in pregnancies complicated by GDM. Further prospective studies are required to enhance the results of this study.

A Prospective, Case-Control Study of Serum Metabolomics in Neonates with Late-Onset Sepsis and Necrotizing Enterocolitis.

Late-onset sepsis (LOS) and necrotizing enterocolitis (NEC) are major causes of neonatal morbidity and mortality. In this prospective, case-control study, we evaluated the metabolic profile of neonates with LOS and NEC. Blood samples were collected from 15 septic neonates and 17 neonates with NEC at the clinical suspicion of the specific diseases. Sixteen gestational and postnatal age-matched neonates without sepsis/NEC served as controls. Serum metabolic profiles were assessed using liquid chromatography-quadrupole time-of-flight mass spectrometry. Metabolomic analysis revealed significant differences in the metabolic profile of neonates with LOS or NEC compared to controls. More specifically, a number of molecules possibly identified as phosphatidylcholines or lysophosphatidylcholines were found to be significantly reduced both in neonates with LOS and those with NEC compared to controls. Additionally, L-carnitine could efficiently discriminate NEC cases from controls. The results of the current study suggest that certain phospholipids and their derivatives could possibly be used as biomarkers for the early detection of LOS and NEC.

Machine Learning Algorithm to Predict Obstructive Coronary Artery Disease: Insights from the CorLipid Trial.

Developing risk assessment tools for CAD prediction remains challenging nowadays. We developed an ML predictive algorithm based on metabolic and clinical data for determining the severity of CAD, as assessed via the SYNTAX score. Analytical methods were developed to determine serum blood levels of specific ceramides, acyl-carnitines, fatty acids, and proteins such as galectin-3, adiponectin, and APOB/APOA1 ratio. Patients were grouped into: obstructive CAD (SS > 0) and non-obstructive CAD (SS = 0). A risk prediction algorithm (boosted ensemble algorithm XGBoost) was developed by combining clinical characteristics with established and novel biomarkers to identify patients at high risk for complex CAD. The study population comprised 958 patients (CorLipid trial (NCT04580173)), with no prior CAD, who underwent coronary angiography. Of them, 533 (55.6%) suffered ACS, 170 (17.7%) presented with NSTEMI, 222 (23.2%) with STEMI, and 141 (14.7%) with unstable angina. Of the total sample, 681 (71%) had obstructive CAD. The algorithm dataset was 73 biochemical parameters and metabolic biomarkers as well as anthropometric and medical history variables. The performance of the XGBoost algorithm had an AUC value of 0.725 (95% CI: 0.691−0.759). Thus, a ML model incorporating clinical features in addition to certain metabolic features can estimate the pre-test likelihood of obstructive CAD.

UHPLC-MS/MS method for the simultaneous determination of nicotine and tobacco-specific nitrosamines NNN and NNK for use in preclinical studies.

A new method was developed and validated for the simultaneous determination of nicotine and tobacco-specific nitrosamines (TSNAs) 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N-nitrosonornicotine (NNN) in two different tests matrices: porcine buccal epithelium tissue and phosphate buffered saline (PBS) extracts of smokeless tobacco products. The novelty of this work is in the development of a liquid chromatography tandem mass spectrometry method that can provide simultaneous quantification of trace levels of TSNAs and high concentrations of nicotine in biological media. Precision, accuracy, and stability were evaluated during method validation to ensure the method was fit for purpose. Several sample preparation and extraction methods were evaluated to minimize matrix effects and maximize analyte recoveries. The method was accurate in the range of 81.1% - 117%; repeatability was estimated in the range of 1.5% - 13.6% across multiple concentrations. The linear regression correlation coefficient (R2) was greater than 0.9959 for all analytes, and the limit of detection (LOD) was determined for nicotine, NNK, and NNN at 1 ng/mL 0.005 ng/mL, and 0.006 ng/ mL, respectively. Our method was found to be appropriate for the analysis of nicotine, NNN, and NNK in the porcine buccal epithelium and PBS extracts of smokeless tobacco products.

Development, Validation and Application of an Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry (UHPLC-MS/MS) Method after QuEChERS Cleanup for Selected Dichloroanilines and Phthalates in Rice Samples.

Dichloroanilines and phthalic acid esters (phthalates) are food contaminants, stable in solution even at high temperatures, which exhibit considerable toxic effects, while acting as endocrine disruptors. In the present study, a quick and easy UHPLC-MS/MS method for simultaneously analyzing two dichloroanilines (3,4-DCA and 3,5-DCA) and six phthalates (DMP, DnBP, BBP, DnOP, DEHP, and mBP) in commercial rice samples was developed, validated, and applied. For the cleanup process, the methodology of quick, easy, cheap, effective, rugged, and safe (QuEChERS) was applied, whereas different dispersants (GCB, C18, and PSA) were tested. What was developed and presented had limits of detection ranging from 0.017 up to 0.12 mg/kg, recoveries (trueness) below 120%, and relative standard deviations (RSD; precision) <15% for all target analytes, whilst no significant matrix effects occurred for all analytes. It was determined that the rice samples analyzed using this developed technique did not contain any of the two dichloroaniline compounds (3,4-DCA and 3,5-DCA) nor two of the six phthalate (DMP and mBP) compounds analyzed, while the levels of other phthalates (DEHP, BBP, DnBP and DnOP) were within the legal limits. The current method ensures a fast and easy approach for the high-throughput quantification of the selected food contaminants in rice.

Correlation of Serum Acylcarnitines with Clinical Presentation and Severity of Coronary Artery Disease.

Recent studies support that acylcarnitines exert a significant role in cardiovascular disease development and progression. The aim of this metabolomics-based study was to investigate the association of serum acylcarnitine levels with coronary artery disease (CAD) severity, as assessed via SYNTAX Score. Within the context of the prospective CorLipid trial (NCT04580173), the levels of 13 circulating acylcarnitines were accurately determined through a newly developed HILIC-MS/MS method in 958 patients undergoing coronary angiography in the AHEPA University Hospital of Thessaloniki, Greece. Patients presenting with acute coronary syndrome had significantly lower median acylcarnitine C8, C10, C16, C18:1 and C18:2 values, compared to patients with chronic coronary syndrome (p = 0.012, 0.007, 0.018, 0.011 and <0.001, respectively). Among CAD subgroups, median C5 levels were significantly decreased in unstable angina compared to STEMI (p = 0.026), while median C10, C16, C18:1 and C18:2 levels were higher in stable angina compared to STEMI (p = 0.019 p = 0.012, p = 0.013 and p < 0.001, respectively). Moreover, median C2, C3, C4 and C8 levels were significantly elevated in patients with diabetes mellitus (p < 0.001, <0.001, 0.029 and 0.011, respectively). Moreover, short-chain acylcarnitine C2, C4, C5 and C6 levels were elevated in patients with heavier calcification and lower left ventricular ejection fraction (LVEF) % (all p-values less than 0.05). With regard to CAD severity, median C4 and C5 levels were elevated and C16 and C18:2 levels were reduced in the high CAD complexity group with SYNTAX Score > 22 (p = 0.002, 0.024, 0.044 and 0.012, respectively), indicating a potential prognostic capability of those metabolites and of the ratio C4/C18:2 for the prediction of CAD severity. In conclusion, serum acylcarnitines could serve as clinically useful biomarkers leading to a more individualized management of patients with CAD, once further clinically oriented metabolomics-based studies provide similar evidence.

Impact of Metabolomics Technologies on the Assessment of Peritoneal Membrane Profiles in Peritoneal Dialysis Patients: A Systematic Review.

Peritoneal dialysis (PD) is an effective and frequent dialysis modality in adults, particularly preferred in infants and young children with end-stage renal disease (ESRD). Long-term exposure of the peritoneal membrane to dialysis solutions results in severe morphologic and functional alterations. Peritoneal dialysis effluent biomarkers are based on omics technologies, which could predict the onset or confirm the diagnosis of peritoneal membrane dysfunction, would allow the development of accurate early prognostic tools and, potentially, the identification of future therapeutic targets. The purpose of our study was to critically review the literature on the impact and the effectiveness of metabolomics technologies in peritoneal health. The main search was performed in electronic databases (PubMed/MEDLINE, Embase and Cochrane Central Register of Controlled Trials) from inception to December 2020, using various combinations of Medical Subject Headings (MeSH). The main search highlighted nine studies, of which seven were evaluated in detail. Metabolomics technologies may provide significant input in the recognition of peritoneal membrane dysfunction in PD patients and provide evidence of early intervention strategies that could protect peritoneum health and function.

Advanced Glycation End-Products (AGEs) of Lysine and Effects of Anti-TCR/Anti-TNF-α Antibody-Based Therapy in the LEW.1AR1-iddm Rat, an Animal Model of Human Type 1 Diabetes.

The LEW.1AR1-iddm rat is an animal model of human type 1 diabetes (T1D). Previously, we have shown that combination with anti-TCR/anti-TNF-α antibody-based therapy re-established normoglycemia and increased proteinic arginine-dimethylation in the spleen, yet not in the pancreas. High blood glucose is often associated with elevated formation of advanced glycation end-products (AGEs) which act via their receptor (RAGE). Both anti-TCR and anti-TNF-α are inhibitors of RAGE. The aim of the present work was to investigate potential biochemical changes of anti-TCR/anti-TNF-α therapy in the LEW.1AR1-iddm rat. We determined by stable-isotope dilution gas chromatography-mass spectrometry (GC-MS) the content of free and proteinic AGEs and the Nε-monomethylation of lysine (Lys) residues in proteins of pancreas, kidney, liver, spleen and lymph nodes of normoglycemic control (ngCo, n = 6), acute diabetic (acT1D, n = 6), chronic diabetic (chT1D, n = 4), and cured (cuT1D, n = 4) rats after anti-TCR/anti-TNF-α therapy. Analyzed biomarkers included Lys and its metabolites Nε-carboxymethyl lysine (CML), furosine and Nε-monomethyl lysine (MML). Other amino acids were also determined. Statistical methods including ANOVA, principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to evaluate the effects. Most statistical differences between the study groups were observed for spleen, pancreas and kidney, with liver and lymph nodes showing no such differences. In the pancreas, the groups differed with respect to proteinic furosine (p = 0.0289) and free CML (p = 0.0023). In the kidneys, the groups differed with respect to proteinic furosine (p = 0.0076) and CML (p = 0.0270). In the spleen, group differences were found for proteinic furosine (p = 0.0114) and free furosine (p = 0.0368), as well as for proteinic CML (p = 0.0502) and proteinic MML (p = 0.0191). The acT1D rats had lower furosine, CML and MML levels in the spleen than the rats in all other groups. This observation corresponds to the lower citrullination levels previously measured in these rats. PCA revealed diametric associations between PC1 and PC2 for spleen (r = -0.8271, p < 0.0001) compared to pancreas (r = 0.5805, p = 0.0073) and kidney (r = 0.8692, p < 0.0001). These findings underscore the importance of the spleen in this animal model of human T1D. OPLS-DA showed that in total sixteen amino acids differed in the experimental groups.

Plasma Lipidomic and Metabolomic Profiling after Birth in Neonates Born to SARS-CoV-19 Infected and Non-Infected Mothers at Delivery: Preliminary Results.

Pregnant women are among the high-risk populations for COVID-19, whereas the risk of vertical transmission to the fetus is very low. Nevertheless, metabolic alternations described in COVID-19 patients may also occur in pregnant women and their offspring. We prospectively evaluated the plasma lipidomic and metabolomic profiles, soon after birth, in neonates born to infected mothers (cases, n = 10) and in the offspring of uninfected ones at delivery (controls, n = 10). All cases had two negative tests for SARS-CoV-2 (nasopharyngeal swabs) performed 72 h apart. Blood samples were obtained within the first hours after birth. Liquid chromatography-high resolution mass spectrometry (UHPLC-TOF/MS) and gas chromatography-mass spectrometry (GC-MS) were applied for the analyses. Multivariate statistical analysis was performed for data evaluation. Changes in several plasma lipid species-classes (long-chain fatty acids phosphatidylcholines, triglycerides), and amino-acids were identified that allowed for clear discrimination between the study groups. The results of this preliminary investigation suggest that neonates born to Sars-Cov-19 positive mothers, without evidence of viral infection at birth, have a distinct plasma lipidomic and metabolomic profile compared to those of uninfected mothers. Whether these findings are reflective of maternal metabolic alternations due to the virus or a metabolic response following an unidentified neonatal infection warrants further investigation.

GC-MS Studies on Derivatization of Creatinine and Creatine by BSTFA and Their Measurement in Human Urine.

In consideration of its relatively constant urinary excretion rate, creatinine (2-amino-1-methyl-5H-imidazol-4-one, MW 113.1) in urine is a useful endogenous biochemical parameter to correct the urinary excretion rate of numerous endogenous and exogenous substances. Reliable measurement of creatinine by gas chromatography (GC)-based methods requires derivatization of its amine and keto groups. Creatinine exists in equilibrium with its open form creatine (methylguanidoacetic acid, MW 131.1), which has a guanidine and a carboxylic group. Trimethylsilylation and trifluoroacetylation of creatinine and creatine are the oldest reported derivatization methods for their GC-mass spectrometry (MS) analysis in human serum using flame- or electron-ionization. We performed GC-MS studies on the derivatization of creatinine (d0-creatinine), [methylo-2H3]creatinine (d3-creatinine, internal standard) and creatine (d0-creatine) with N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) using standard derivatization conditions (60 min, 60 °C), yet in the absence of any base. Reaction products were characterized both in the negative-ion chemical ionization (NICI) and in the positive-ion chemical ionization (PICI) mode. Creatinine and creatine reacted with BSTFA to form several derivatives. Their early eluting N,N,O-tris(trimethylsilyl) derivatives (8.9 min) were found to be useful for the precise and accurate measurement of the sum of creatinine and creatine in human urine (10 µL, up to 20 mM) by selected-ion monitoring (SIM) of m/z 271 (d0-creatinine/d0-creatine) and m/z 274 (d3-creatinine) in the NICI mode. In the PICI mode, SIM of m/z 256, m/z 259, m/z 272 and m/z 275 was performed. BSTFA derivatization of d0-creatine from a freshly prepared solution in distilled water resulted in formation of two lMate-eluting derivatives (14.08 min, 14.72 min), presumably creatinyl-creatinine, with the creatininyl residue existing in its enol form (14.08 min) and keto form (14.72 min). Our results suggest that BSTFA derivatization does not allow specific analysis of creatine and creatinine by GC-MS. Preliminary analyses suggest that pentafluoropropionic anhydride (PFPA) is also not useful for the measurement of creatinine in the presence of creatine. Both BSTFA and PFPA facilitate the conversion of creatine to creatinine. Specific measurement of creatinine in urine is possible by using pentafluorobenzyl bromide in aqueous acetone.