Predicting effect of anti-PD-1/PD-L1 inhibitors therapy for hepatocellular carcinoma by detecting plasma metabolite based on UHPLC-MS.

Introduction: Anti-PD-1/PD-L1 inhibitors therapy has become a promising treatment for hepatocellular carcinoma (HCC), while the therapeutic efficacy varies significantly among effects for individual patients are significant difference. Unfortunately, specific predictive biomarkers indicating the degree of benefit for patients and thus guiding the selection of suitable candidates for immune therapy remain elusive.no specific predictive biomarkers are available indicating the degree of benefit for patients and thus screening the preferred population suitable for the immune therapy. Methods: Ultra-high-pressure liquid chromatography-mass spectrometry (UHPLC-MS) considered is an important method for analyzing biological samples, since it has the advantages of high rapid, high sensitivity, and high specificity. Ultra-high-pressure liquid chromatography-mass spectrometry (UHPLC-MS) has emerged as a pivotal method for analyzing biological samples due to its inherent advantages of rapidity, sensitivity, and specificity. In this study, potential metabolite biomarkers that can predict the therapeutic effect of HCC patients receiving immune therapy were identified by UHPLC-MS. Results: A partial least-squares discriminant analysis (PLS-DA) model was established using 14 glycerophospholipid metabolites mentioned above, and good prediction parameters (R2 = 0.823, Q2 = 0.615, prediction accuracy = 0.880 and p < 0.001) were obtained. The relative abundance of glycerophospholipid metabolite ions is closely related to the survival benefit of HCC patients who received immune therapy. Discussion: This study reveals that glycerophospholipid metabolites play a crucial role in predicting the efficacy of immune therapy for HCC.

Targeted metabolomics-based understanding of the sleep disturbances in drug-naïve patients with schizophrenia.

BACKGROUND: Sleep disturbances are a common occurrence in patients with schizophrenia, yet the underlying pathogenesis remain poorly understood. Here, we performed a targeted metabolomics-based approach to explore the potential biological mechanisms contributing to sleep disturbances in schizophrenia. METHODS: Plasma samples from 59 drug-naïve patients with schizophrenia and 36 healthy controls were subjected to liquid chromatography-mass spectrometry (LC-MS) targeted metabolomics analysis, allowing for the quantification and profiling of 271 metabolites. Sleep quality and clinical symptoms were assessed using the Pittsburgh Sleep Quality Index (PSQI) and the Positive and Negative Symptom Scale (PANSS), respectively. Partial correlation analysis and orthogonal partial least squares discriminant analysis (OPLS-DA) model were used to identify metabolites specifically associated with sleep disturbances in drug-naïve schizophrenia. RESULTS: 16 characteristic metabolites were observed significantly associated with sleep disturbances in drug-naïve patients with schizophrenia. Furthermore, the glycerophospholipid metabolism (Impact: 0.138, p<0.001), the butanoate metabolism (Impact: 0.032, p=0.008), and the sphingolipid metabolism (Impact: 0.270, p=0.104) were identified as metabolic pathways associated with sleep disturbances in drug-naïve patients with schizophrenia. CONCLUSIONS: Our study identified 16 characteristic metabolites (mainly lipids) and 3 metabolic pathways related to sleep disturbances in drug-naïve schizophrenia. The detection of these distinct metabolites provide valuable insights into the underlying biological mechanisms associated with sleep disturbances in schizophrenia.

Feasibility of a telehealth-based contingency management intervention for alcohol use disorders using the phosphatidylethanol (PEth) 16:0/18:1 alcohol biomarker: a pilot randomized trial.

Background: Phosphatidylethanol (PEth) is a blood-based biomarker for alcohol consumption that can be self-collected and has high sensitivity, specificity, and a longer detection window compared to other alcohol biomarkers.Objectives: We evaluated the feasibility and acceptability of a telehealth-based contingency management (CM) intervention for alcohol use disorder (AUD) using the blood-based biomarker PEth to assess alcohol consumption.Methods: Sixteen adults (7 female, 9 male) with AUD were randomized to Control or CM conditions. Control participants received reinforcers regardless of their PEth levels. CM participants received reinforcers for week-to-week decreases in PEth (Phase 1) or maintenance of PEth consistent with abstinence (<20 ng/mL, Phase 2). Blood samples were self-collected using the TASSO-M20 device. Acceptability was assessed by retention in weeks. Satisfaction was assessed with the Client Satisfaction Questionnaire (CSQ-8) and qualitative interviews. The primary efficacy outcome was PEth-defined abstinence. Secondary outcomes included the proportion of visits with PEth-defined heavy alcohol consumption, negative urine ethyl glucuronide results, and self-reported alcohol use.Results: Retention averaged 18.6 ± 8.8 weeks for CM participants. CM participants reported high levels of satisfaction (CSQ-8, Mean = 30.3 ± 1.5). Interview themes included intervention positives, such as staff support, quality of life improvement, and accountability. 72% of PEth samples from CM participants were consistent with abstinence versus 34% for Control participants (OR = 5.0, p = 0.007). PEth-defined heavy alcohol consumption was detected in 28% of CM samples and 52% of Control samples (OR = 0.36, p = 0.159). CM participants averaged 1.9 ± 1.7 drinks/day versus 4.2 ± 6.3 for Control participants (p = 0.304).Conclusion: Results support the acceptability and satisfaction of a telehealth PEth-based CM intervention, though a larger study is needed to assess its efficacy [NCT04038021].

Significantly elevated phosphatidylethanol levels in recent suicide attempters, but not in depressed controls and healthy volunteers.

INTRODUCTION: Suicide is a complex transdiagnostic phenomenon. It is strongly associated with, but not exclusive to major depressive disorder (MDD). Hazardous alcohol drinking has also been linked to an increased risk of suicidal behaviours, however, it is often underreported. The study aimed to evaluate whether an objective measure of chronic alcohol use, phosphatidylethanol (PEth) could be useful as a biomarker in clinical practice. METHOD: ology. The present case-control multi-centric study recruited 156 participants into three study groups: 52 patients treated for major depressive disorder (MDD), 51 individuals immediately following a suicide attempt (SA), and 53 volunteers. Sociodemographic data, medical history, and laboratory data, including PEth concentrations and C-reactive protein levels, were collected from study participants. RESULTS: PEth concentrations were the highest in suicide attempters (232,54 ± 394,01 ng/ml), followed by patients with MDD (58,39 ± 135,82 ng/ml), and the control group (24,45 ± 70,83 ng/ml) (Kruskall Wallis χ2 = 12.23, df = 2, p = .002). In a multinomial logistic regression model with adjustments, PEth concentration was able to predict belonging to suicide attempters' group, but not to depression group (p = .01). Suicide attempters were also more likely to underreport their recent alcohol consumption. LIMITATIONS: We did not analyze SA methods, psychiatric comorbidity and several other factors that might be associated with PEth levels, such as body mass index, race, and haemoglobin levels. Sample recruited in hospital settings may not be representative of the whole population. The results of this adult-only study cannot be generalized to adolescents. CONCLUSIONS: PEth levels in recent suicide attempters significantly exceeded those of patients with MDD and controls. Suicide attempters also were more likely to underreport their alcohol consumption when questioned about their consuption. PEth might be an interesting biomarker to evaluate individuals at risk of SA.

Liquid chromatography-mass spectrometry method for discovering the metabolic markers to reveal the potential therapeutic effects of naringin on osteoporosis.

The pharmacological effects and therapeutic targets of naringin (NG) against osteoporosis (OP) is still unclear. Liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-TOF/MS) based non-targeted metabonomics has been used to explore the differentiated metabolites and potential biological pathways of NG in the pathological process of OP. Using network pharmacology analysis, the key protein targets of NG against OP were also screened. By the metabonomics analysis, a total of 33 differentiated metabolites in serum were discovered, of which 21 were significantly regulated by NG treatment. These metabolites majorly associated with to amino acid metabolism,polyunsaturated fatty acid metabolism, pyruvate metabolism and glycerophospholipidmetabolism. Using the network pharmacology prediction analysis, NG was related to the expression changes of 13 important protein targets. It showed that high-throughput metabonomics strategy integrated with network pharmacology could insight into molecular mechanisms of natural products.

Validation of a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for erythrocyte phosphatidylethanol revealing critical considerations for its use as a clinical biomarker.

Erythrocyte membrane-incorporated phosphatidylethanol (PEth) forms only in the presence of ethanol and, once formed, provides a persisting marker for historical alcohol consumption. Relationships between PEth concentration, extent of consumption and time from consumption are under investigation. Threshold values of PEth have been proposed as indicators for any, or for harmful alcohol consumption. Here, we describe an assay for erythrocyte PEth 16:0/18:1 that offers the efficiency needed for routine clinical deployment, in the context of a fully validated methodology. However, we observe that conventional procedures for validating assay methodology are insufficient where the analyte of interest, membrane-incorporated PEth 16:0/18:1, has different physicochemical properties to the soluble PEth 16:0/18:1 and PEth 16:0/18:1-d5 that are used for making calibrator, controls and internal standards. Whereas the internal standard did fully correct for differences in matrix effects and recovery when different extraction solvents were applied to calibrators and controls (in soluble form), it failed to correct for a 1.5-fold difference in the relative efficiency of two solvents, in this case, acetonitrile and isopropanol in extracting PEth from erythrocyte membrane in clinical samples. Differences in the efficiency of the extraction of membrane-bound PEth translate to different results from the same specimen. That can mean that threshold values derived by one methodology cannot be safely generalised to another. That hampers the generalisability of individual laboratory's experience with PEth assay results. Harmonising extraction methodology between laboratories becomes very important where membrane-incorporated PEth itself remains unavailable as an assay standard.

Development and validation of an analytical method for the simultaneous determination of the alcohol biomarkers ethyl glucuronide, ethyl sulfate, N-acetyltaurine, and 16:0/18:1-phosphatidylethanol in human blood.

As alcohol is the most common addictive substance worldwide, it is inevitable to advance the established research. New and more substantial analytical methods can be applied to reply to complex questions in legal or forensic contexts. Therefore, an analytical method for the simultaneous determination of four different alcohol biomarkers-ethyl glucuronide, ethyl sulfate, N-acetyltaurine, and 16:0/18:1-phosphatidylethanol-in human blood was developed, validated, and verified. Despite the different chemical properties of the analytes, a specific determination via HPLC-MS/MS was achieved using a novel type of a Phenomenex Luna® Omega Sugar column. Furthermore, all criteria for a successful validation were fulfilled according to forensic guidelines. The method proved to be linear and demonstrates selectivity and sufficient sensitivity for every biomarker. LODs obtained with this method of 2.6 ng/ml (EtG), 4.7 ng/ml (EtS), 12.5 ng/ml (NAcT), and 6.9 ng/ml (PEth) were in an acceptable range for routine applications, and the stability of all analytes over a range of 12 h is given. The verification of the new developed method was performed with authentic samples. Thus, whole blood and postmortem samples were analyzed to obtain information about the drinking behavior, which can answer complex questions regarding alcohol consumption.

Associations of plasma glycerophospholipid profile with modifiable lifestyles and incident diabetes in middle-aged and older Chinese.

AIMS/HYPOTHESIS: Glycerophospholipid (GPL) perturbance was linked to the pathogenesis of diabetes in animal studies but prospective studies in humans are rare, particularly in Asians. We aimed to investigate the associations between plasma GPLs and incident diabetes and to explore effects of lifestyle on the associations in a Chinese population. METHODS: The study included 1877 community-dwelling Chinese individuals aged 50-70 years (751 men and 1126 women), free of diabetes at baseline and followed for 6 years. A total of 160 GPL species were quantified in plasma at baseline by using high-throughput targeted lipidomics. Log-Poisson regression was used to assess the associations between GPLs and incidence of diabetes. RESULTS: Over the 6 years of follow-up, 499 participants (26.6%) developed diabetes. After multivariable adjustment, eight GPLs were positively associated with incident diabetes (RRper SD 1.13-1.25; all false-discovery rate [FDR]-corrected p < 0.05), including five novel GLPs, namely phosphatidylcholines (PCs; 16:0/18:1, 18:0/16:1, 18:1/20:3), lysophosphatidylcholine (LPC; 20:3) and phosphatidylethanolamine (PE; 16:0/16:1), and three reported GPLs (PCs 16:0/16:1, 16:0/20:3 and 18:0/20:3). In network analysis, a PC-containing module was positively associated with incident diabetes (RRper SD 1.16 [95% CI 1.06, 1.26]; FDR-corrected p < 0.05). Notably, three of the diabetes-associated PCs (16:0/16:1, 16:0/18:1 and 18:0/16:1) and PE (16:0/16:1) were associated not only with fatty acids in the de novo lipogenesis (DNL) pathway, especially 16:1n-7 (Spearman correlation coefficients = 0.35-0.62, p < 0.001), but also with an unhealthy dietary pattern high in refined grains and low in fish, dairy and soy products (|factor loadings| ≥0.2). When stratified by physical activity levels, the associations of the eight GPLs and the PC module with incident diabetes were stronger in participants with lower physical activity (RRper SD 1.24-1.49, FDR-corrected p < 0.05) than in those with the median and higher physical activity levels (RRper SD 1.03-1.12, FDR-corrected p ≥ 0.05; FDR-corrected pinteraction < 0.05). CONCLUSIONS/INTERPRETATION: Eight GPLs, especially PCs associated with the DNL pathway, were positively associated with incident diabetes in a cohort of Chinese men and women. The associations were most prominent in participants with a low level of physical activity.

Quantitation of phosphatidylethanols in dried blood spots to determine rates of prenatal alcohol exposure in Ontario.

BACKGROUND: Estimating rates of prenatal alcohol exposure (PAE) in a population is necessary to ensure that proper medical and social supports and interventions are in place. This study sought to estimate PAE in Ontario, Canada by quantifying phosphatidylethanol (PEth) homologues in over 2000 residual neonatal dried blood spots (DBS). METHODS: A random selection of 2011 residual DBS collected over a 1-week time period were anonymized and extracted. A targeted liquid chromatography-mass spectrometry method was used to quantify 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanol (PEth (16:0/18:1) or POPEth), the clinically accepted biomarker, and six additional PEth homologues. A POPEth level above the United States Drug Testing Laboratories (USDTL) cutoff up to 4 weeks predelivery was indicative of PAE. All PEth homologues were correlated to one another and logistic regression was used to determine the association between PAE status and infant characteristics. RESULTS: The estimated rate of PAE in Ontario, up to the last 4 weeks of gestation, was 15.5% (POPEth >28.5 nM). Most PEth homologues were moderately to strongly correlated to one another. A low birth weight and preterm birth were both associated with PAE, while being small for gestational age had lower odds of PAE. CONCLUSIONS: The results of this study suggest that PAE may be more prevalent in Ontario than previous estimates by self-report or meconium testing. These findings support the need to consider the effectiveness of current interventions and the design of new interventions to address this significant public health issue.

The alcohol marker phosphatidylethanol is closely related to AST, GGT, ferritin and HDL-C.

BACKGROUND: The aim of this study was to evaluate the quantitative relation between common clinical chemical analyses and ethanol use, measured by a combination of the two alcohol markers phosphatidylethanol (PEth) and carbohydrate-deficient transferrin (CDT). METHODS: Results of PEth and CDT in whole blood and serum, respectively, were included, together with information on 10 different commonly measured clinical chemical analytes, as well as age and sex. PEth was analysed by UPC2 -MS/MS and CDT was measured by capillary electrophoresis. RESULTS: Samples from 4873 patients were included. The strongest relation to alcohol consumption as measured by PEth, when correcting for age and sex, was found for HDL-C (standardized ß = 0.472, p < 0.001), AST (standardized ß = 0.372, p < 0.001), ferritin (standardized ß = 0.332, p < 0.001) and GGT (standardized ß = 0.325, p < 0.001). The relation to PEth was weak for total cholesterol, TG and ALP. No relation was found for Hb and LDL-C. CONCLUSIONS: When using PEth as a marker for alcohol consumption, this study demonstrated the quantitative relation to commonly used test as AST or GGT, but also an important relation to ferritin or HDL-C. In clinical practice, elevated levels of these clinical chemical analytes should initiate further work-up on possibly harmful alcohol use.

Alcohol use and dysglycemia among people living with human immunodeficiency virus (HIV) in the Alcohol & Metabolic Comorbidities in PLWH: Evidence Driven Interventions (ALIVE-Ex) study.

BACKGROUND: At-risk alcohol use is a common and costly form of substance misuse that is highly prevalent among people living with HIV (PLWH). The goal of the current analysis was to test the hypothesis that PLWH with at-risk alcohol use are more likely to meet the clinical criteria for prediabetes/diabetes than PLWH with low-risk alcohol use. METHODS: A cross-sectional analysis was performed on measures of alcohol and glycemic control in adult PLWH (n = 105) enrolled in a prospective, interventional study (the ALIVE-Ex Study (NCT03299205)) that investigated the effects of aerobic exercise on metabolic dysregulation in PLWH with at-risk alcohol use. The Alcohol Use Disorders Identification Test (AUDIT), Timeline Followback, and phosphatidylethanol (PEth) level were used to measure alcohol use. Participants were stratified into low-risk (AUDIT score < 5) and at-risk alcohol use (AUDIT  score ≥ 5). All participants underwent an oral glucose tolerance test and measures of glycemic control- the Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) and Matsuda Index - were correlated with alcohol measures and compared by AUDIT score group using mixed-effects linear and logistic regression models, adjusting for age, sex, race, body mass index (BMI), and viral load. RESULTS: In response to the glucose challenge, participants with at-risk alcohol use (n = 46) had higher glucose levels and were five times more likely to meet criteria for prediabetes/diabetes (OR: 5.3 (1.8, 15.9)) than participants with an AUDIT score < 5. Two-hour glucose values were positively associated with AUDIT score and PEth level and a higher percentage of PLWH with at-risk alcohol use had glucose values ≥140 mg/dl than those with low-risk alcohol use (34.8% vs. 10.2%, respectively). CONCLUSION: In this cohort of PLWH, at-risk alcohol use increased the likelihood of meeting the clinical criteria for prediabetes/diabetes (2-h glucose level ≥140 mg/dl). Established determinants of metabolic dysfunction (e.g., BMI, waist-hip ratio) were not associated with greater alcohol use and dysglycemia, suggesting that other mechanisms may contribute to the impaired glycemic control observed in this cohort.

Objective assessment of alcohol consumption in early pregnancy using phosphatidylethanol: a cross-sectional study.

BACKGROUND: Alcohol consumption during pregnancy is associated with major birth defects and developmental disabilities. Questionnaires concerning alcohol consumption during pregnancy underestimate alcohol use while the use of a reliable and objective biomarker for alcohol consumption enables more accurate screening. Phosphatidylethanol can detect low levels of alcohol consumption in the previous two weeks. In this study we aimed to biochemically assess the prevalence of alcohol consumption during early pregnancy using phosphatidylethanol in blood and compare this with self-reported alcohol consumption. METHODS: To evaluate biochemically assessed prevalence of alcohol consumption during early pregnancy using phosphatidylethanol levels, we conducted a prospective, cross-sectional, single center study in the largest tertiary hospital of the Netherlands. All adult pregnant women who were under the care of the obstetric department of the Erasmus MC and who underwent routine blood testing at a gestational age of less than 15 weeks were eligible. No specified informed consent was needed. RESULTS: The study was conducted between September 2016 and October 2017. In total, we received 1,002 residual samples of 992 women. After applying in- and exclusion criteria we analyzed 684 samples. Mean gestational age of all included women was 10.3 weeks (SD 1.9). Of these women, 36 (5.3 %) tested positive for phosphatidylethanol, indicating alcohol consumption in the previous two weeks. Of women with a positive phosphatidylethanol test, 89 % (n = 32) did not express alcohol consumption to their obstetric care provider. CONCLUSIONS: One in nineteen women consumed alcohol during early pregnancy with a high percentage not reporting this use to their obstetric care provider. Questioning alcohol consumption by an obstetric care provider did not successfully identify (hazardous) alcohol consumption. Routine screening with phosphatidylethanol in maternal blood can be of added value to identify women who consume alcohol during pregnancy.

Factors associated with phosphatidylethanol (PEth) sensitivity for detecting unhealthy alcohol use: An individual patient data meta-analysis.

BACKGROUND: Objective measurement of alcohol consumption is important for clinical care and research. Adjusting for self-reported alcohol use, we conducted an individual participant data (IPD) meta-analysis to examine factors associated with the sensitivity of phosphatidylethanol (PEth), an alcohol metabolite, among persons self-reporting unhealthy alcohol consumption. METHODS: We identified 21 eligible studies and obtained 4073 observations from 3085 participants with Alcohol Use Disorders Identification Test-Consumption (AUDIT-C) positive scores (≥3 for women and ≥4 for men) and PEth measurements. We conducted 1-step IPD meta-analysis using mixed effects models with random intercepts for study site. We examined the associations between demographic (sex, race/ethnicity, and age) and biologic (body mass index-BMI, hemoglobin, HIV status, liver fibrosis, and venous versus finger-prick blood collection) variables with PEth sensitivity (PEth≥8 ng/ml), adjusting for the level of self-reported alcohol use using the AUDIT-C score. RESULTS: One third (31%) of participants were women, 32% were African, 28% African American, 28% White, and 12% other race/ethnicity. PEth sensitivity (i.e., ≥8 ng/ml) was 81.8%. After adjusting for AUDIT-C, we found no associations of sex, age, race/ethnicity, or method of blood collection with PEth sensitivity. In models that additionally included biologic variables, those with higher hemoglobin and indeterminate and advanced liver fibrosis had significantly higher odds of PEth sensitivity; those with higher BMI and those living with HIV had significantly lower odds of PEth sensitivity. African Americans and Africans had higher odds of PEth sensitivity than whites in models that included biologic variables. CONCLUSIONS: Among people reporting unhealthy alcohol use, several biological factors (hemoglobin, BMI, liver fibrosis, and HIV status) were associated with PEth sensitivity. Race/ethnicity was associated with PEth sensitivity in some models but age, sex, and method of blood collection were not. Clinicians should be aware of these factors, and researchers should consider adjusting analyses for these characteristics where possible.

Plasma glycerophospholipid profile, erythrocyte n-3 PUFAs, and metabolic syndrome incidence: a prospective study in Chinese men and women.

BACKGROUND: Animal studies have highlighted critical roles of glycerophospholipid (GP) metabolism in various metabolic syndrome (MetS)-related features such as dyslipidemia, obesity, and insulin resistance. However, human prospective studies of associations between circulating GPs and risks of MetS are scarce. OBJECTIVES: We aimed to investigate whether GPs are associated with incidence of MetS in a well-established cohort. METHODS: A total of 1243 community-dwelling Chinese aged 50-70 y without MetS at baseline and followed up for 6 y were included in current analyses. A total of 145 plasma GPs were quantified by high-throughput targeted lipidomics. MetS was defined using the updated National Cholesterol Education Program Adult Treatment Panel III criteria for Asian Americans. RESULTS: After 6 y, 429 participants developed MetS. Eleven GPs, especially those with long-chain polyunsaturated fatty acids (LCPUFAs) or very-long-chain polyunsaturated fatty acids (VLCPUFAs) at the sn-2 position, including 1 phosphatidylcholine (PC) [PC(18:0/22:6)], 9 phosphatidylethanolamines (PEs) [PE(16:0/22:6), PE(18:0/14:0), PE(18:0/18:1), PE(18:0/18:2), PE(18:0/20:3), PE(18:0/22:5), PE(18:0/22:6), PE(18:1/22:6), and PE(18:2/22:6)], and 1 phosphatidylserine (PS) [PS(18:0/18:0)], were positively associated with incident MetS (RRs: 1.16-1.30 per SD change; Bonferroni-corrected P < 0.05). In network analysis, the strongest positive association for MetS incidence was evidenced in a module mainly composed of PEs containing C22:6 and PSs [RR: 1.21; 95% CI: 1.12, 1.31 per SD change; Bonferroni-corrected P < 0.05]. This association was more pronounced in participants with lower erythrocyte total n-3 PUFA concentrations [Bonferroni-corrected Pinter(P value for the interaction)< 0.05]. CONCLUSIONS: Elevated plasma concentrations of GPs, especially PEs with LCPUFAs or VLCPUFAs at the sn-2 position, are associated with higher risk of incident MetS. Future studies are merited to confirm our findings.

Screening of lipid metabolism biomarkers in patients with coronary heart disease via ultra-performance liquid chromatography-high resolution mass spectrometry.

Coronary heart disease (CHD) has a high mortality worldwide. This study aimed to screen lipid metabolism biomarkers in patients with coronary heart disease via ultra-performance liquid chromatography-high resolution mass spectrometry. Extraction and reconstitution solvents, liquid chromatographic and mass spectrometry conditions were optimized to detect more plasma lipid metabolites. In this study, the chromatographic and mass spectra characteristics of lipid metabolites were summarized. A total of 316 lipid metabolites were annotated via diagnostic fragment ion filtration, nitrogen rule filtration, and neutral loss filtration. Glycerophospholipid metabolism and sphingolipid metabolism were revealed as the main lipid disorders of CHD. This study provides a novel insight for high-throughput detection of lipid metabolites in plasma and provides a further understanding of the occurrence of CHD, which can provide valuable suggestions for the prevention of CHD.

The Association between the Alcohol Biomarker Phosphatidylethanol (PEth) and Self-Reported Alcohol Consumption among Russian and Norwegian Medical Patients.

AIMS: Valid measures to identify harmful alcohol use are important. Alcohol Use Disorders Identification Test (AUDIT) is a validated questionnaire used to self-report harmful drinking in several cultures and settings. Phosphatidylethanol 16:0/18:1 (PEth) is a direct alcohol biomarker measuring alcohol consumption levels. The aim of this study was to investigate how PEth levels correlate with AUDIT-QF and weekly grams of alcohol consumed among patients in two urban hospitals. In addition, we wanted to investigate the predictive value of PEth in identifying harmful alcohol use as defined by AUDIT-QF and weekly grams of alcohol cutoffs. METHODS: A cross-sectional study comprising acute medically ill patients with measurable PEth levels (≥0.030 µM) admitted to two urban hospitals in Oslo, Norway (N = 931) and Moscow, Russia (N = 953) was conducted using PEth concentrations in whole blood, sociodemographic data and AUDIT-QF questionnaires. RESULTS: PEth levels from patients with measurable PEth were found to be positively correlated with AUDIT-QF scores, with PEth cutpoints of 0.128 µM (Oslo) and 0.270 µM (Moscow) providing optimal discrimination for harmful alcohol use defined by AUDIT-QF (the difference between cities probably reflecting different national drinking patterns in QF). When converting AUDIT-QF into weekly grams of alcohol consumed, the predictive value of PEth improved, with optimal PEth cutpoints of 0.327 (Oslo) and 0.396 (Moscow) µM discriminating between harmful and non-harmful alcohol use as defined in grams (≥350 grams/week). CONCLUSIONS: By using PEth levels and converting AUDIT-QF into weekly grams of alcohol it was possible to get an improved rapid and sensitive determination of harmful alcohol use among hospitalized patients.

Measurement of the alcohol biomarker phosphatidylethanol (PEth) in dried blood spots and venous blood-importance of inhibition of post-sampling formation from ethanol.

Phosphatidylethanol (PEth) is a group of phospholipids formed in cell membranes following alcohol consumption by action of the enzyme phospholipase D (PLD). PEth measurement in whole blood samples is established as a specific alcohol biomarker with clinical and forensic applications. However, in blood specimens containing ethanol, formation of PEth may continue after sampling leading to falsely elevated concentrations. This study evaluated the use of dried blood spot (DBS) and microsampling specimens to avoid post-sampling formation of PEth. Filter paper cards and three commercial devices for volumetric microsampling of finger-pricked blood were assessed, using PEth-negative and PEth-positive whole blood fortified with 2 g/L ethanol. PEth (16:0/18:1) was measured by LC-MS/MS. Post-sampling formation of PEth occurred in wet blood and in the volumetric devices, but not filter paper cards, when stored at room temperature for 48 h. Addition of an inhibitor of PLD, sodium metavanadate (NaVO3), eliminated post-sampling formation during storage and drying. In conclusion, the present study confirmed previous observations that PEth can be formed in blood samples after collection, if the specimen contains ethanol. The results further demonstrated that post-sampling formation of PEth from ethanol also occurred with commercial devices for volumetric dried blood microsampling. In order for a PEth result not to be questioned, it is recommended to use a PLD inhibitor, whether venous blood is collected in a vacutainer tube or finger-pricked blood is obtained using devices for dried blood microsampling.

Phosphatidylethanol in whole blood of rhesus monkeys correlates with ethanol consumption.

BACKGROUND: Phosphatidylethanol (PEth) homologs are ethanol metabolites used to identify and monitor alcohol drinking in humans. In this study, we measured levels of the 2 most abundant homologs, PEth 16:0/18:1 and PEth 16:0/18:2, in whole blood samples from rhesus macaque monkeys that drank ethanol daily ad libitum to assess the relationship between PEth levels and recent ethanol exposure in this animal model. METHODS: Blood samples were obtained from The Monkey Alcohol Tissue Research Resource. The monkeys were first induced to consume 4% (w/v) ethanol in water from a panel attached to their home cage. Then, monkeys were allowed to drink ethanol and water ad libitum 22 h daily for 12 months and the daily amount of ethanol each monkey consumed was measured. Whole, uncoagulated blood was collected from each animal at the end of the entire experimental procedure. PEth 16:0/18:1 and PEth 16:0/18:2 levels were analyzed by HPLC with tandem mass spectrometry, and the ethanol consumed during the preceding 14 days was measured. Combined PEth was the sum of the concentrations of both homologs. RESULTS: Our results show that (1) PEth accumulates in the blood of rhesus monkeys after ethanol consumption; (2) PEth homolog levels were correlated with the daily average ethanol intake during the 14-day period immediately preceding blood collection; (3) the application of established human PEth 16:0/18:1 cutoff concentrations indicative of light social or no ethanol consumption (<20 ng/ml), moderate ethanol consumption (≥ 20 and < 200 ng/ml) and heavy ethanol consumption (≥ 200 ng/ml) predicted significantly different ethanol intake in these animals. PEth homologs were not detected in ethanol-naïve controls. CONCLUSIONS: This study confirms that PEth is a sensitive biomarker for ethanol consumption in rhesus macaque monkeys. This nonhuman primate model may prove useful in evaluating sources of variability previously shown to exist between ethanol consumption and PEth homolog levels among humans.

Phosphatidylethanol as Blood Biomarker of Alcohol Consumption in Early Pregnancy: An Observational Study in 4,067 Pregnant Women.

BACKGROUND: The teratogenic effects of alcohol are well documented, but there is a lack of screening methods to detect alcohol use during pregnancy. Phosphatidylethanol 16:0/18:1 (PEth) is a specific and sensitive biomarker reflecting alcohol intake up to several weeks after consumption. The aim of this study was to investigate the prevalence of positive PEth values as an indicator of early prenatal alcohol exposure in a general population of pregnant women. METHODS: Rhesus typing is routinely performed in Norway in all pregnancies around gestational week 12. Rhesus-negative women have an additional test taken around week 24. Blood samples submitted to St. Olav University Hospital in Trøndelag, Norway, for Rhesus typing during the period September 2017 to October 2018 were collected. A total of 4,533 whole blood samples from 4,067 women were analyzed for PEth (limit of quantification of 0.003 µM). RESULTS: Fifty-eight women had a positive PEth sample. Of these, 50 women were positive around gestational week 12, 3 women were positive around week 24, and in 5 cases, the timing was unknown. There were no significant differences in proportions of women with positive PEth values related to age, or rural versus urban residency. CONCLUSION: In an unselected pregnant population in Norway, 1.4% had a positive PEth sample around gestational week 12, whereas 0.4% had a positive sample around week 24. The use of PEth as an alcohol biomarker should be further investigated as a diagnostic tool in the antenatal setting.

Stability of phosphatidylethanol 16:0/18:1 in authentic and spiked whole blood.

Phosphatidylethanol 16:0/18:1 (PEth) is the most abundant homologue of the phosphatidylethanol group of phospholipids. Formed only in the presence of ethanol, PEth is used as a biomarker in whole blood to provide information about the consumption of alcohol. As information on the storage life of PEth is essential for its beneficial use as a biomarker, this study investigated the stability of PEth in spiked and authentic whole blood samples stored at 4°C. Human whole blood samples (n = 23) and spiked whole blood samples (n = 7) with a concentration range between 5 and 2000 ng/ml were analysed at specific time intervals, up to 90 days. Differences were evident between the stability of authentic and spiked samples. PEth was stable at 4°C for 60 days (concentrations within 15% of initial concentration) in authentic samples, whereas spiked samples were stable for up to 30 days. This study emphasizes the importance of including authentic samples in stability experiments.