Urinary antihypertensive drug metabolite screening using molecular networking coupled to high-resolution mass spectrometry fragmentation

Introduction

Mass spectrometry is the current technique of choice in studying drug metabolism. High-resolution mass spectrometry in combination with MS/MS gas-phase experiments has the potential to contribute to rapid advances in this field. However, the data emerging from such fragmentation spectral files pose challenges to downstream analysis, given their complexity and size.

Objectives

This study aims to detect and visualize antihypertensive drug metabolites in untargeted metabolomics experiments based on the spectral similarity of their fragmentation spectra. Furthermore, spectral clusters of endogenous metabolites were also examined.

Methods

Here we apply a molecular networking approach to seek drugs and their metabolites, in fragmentation spectra from urine derived from a cohort of 26 patients on antihypertensive therapy. The mass spectrometry data was collected on a Thermo Q-Exactive coupled to pHILIC chromatography using data dependent analysis (DDA) MS/MS gas-phase experiments.

Results

In total, 165 separate drug metabolites were found and structurally annotated (17 by spectral matching and 122 by classification based on a clustered fragmentation pattern). The clusters could be traced to 13 drugs including the known antihypertensives verapamil, losartan and amlodipine. The molecular networking approach also generated clusters of endogenous metabolites, including carnitine derivatives, and conjugates containing glutamine, glutamate and trigonelline.

Conclusions

The approach offers unprecedented capability in the untargeted identification of drugs and their metabolites at the population level and has great potential to contribute to understanding stratified responses to drugs where differences in drug metabolism may determine treatment outcome.

     

Quantitative analysis of tetrahydrofolate metabolites from <Emphasis Type="Italic">clostridium autoethanogenum</Emphasis>

Introduction

Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.

Objective

To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.

Methods

Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.

Results

Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.

Conclusion

Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.

     

Urine metabolic fingerprinting can be used to predict the risk of metritis and highlight the pathobiology of the disease in dairy cows

Introduction

Metritis is an uterine pathology that causes economic losses for the dairy industry. It is associated with lower reproductive efficiency, increased culling rates, decreased milk production and increased veterinary costs.

Objectives

To gain a more detailed view of the urine metabolome and to detect metabolite signature in cows with metritis. In addition, we aimed to identify early metabolites which can help to detect cows at risk to develop metritis in the future.

Methods

We used nuclear magnetic resonance spectroscopy starting at 8 and 4 weeks prior to the expected day of parturition, during the week of diagnosis of metritis, and at 4 and 8 weeks after diagnosis of metritis in Holstein dairy cows.

Results

At 8 weeks before parturition, pre-metritic cows had a total of 30 altered metabolites. Interestingly, 28 of them increased in urine when compared with control cows (P?<?0.05). At 4 weeks before parturition, 34 metabolites were altered. At the week of diagnosis of metritis a total of 20 metabolites were altered (P?<?0.05). The alteration continued at 4 and 8 weeks after diagnosis.

Conclusions

The metabolic fingerprints in the urine of pre-metritic and metritic cows point toward excretion of multiple amino acids, tricarboxylic acid cycle metabolites and monosaccharides. Combination of galactose, leucine, lysine and panthotenate at 8 weeks before parturition might serve as predictive biomarkers for metritis.

     

Extraction of natural products from bark of <Emphasis Type="Italic">Betula pendula</Emphasis> using ionic liquids

Introduction

Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.

Objectives

To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.

Methods

The extracted compounds were analyzed by LC/MS and GC/MS.

Results

The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.

Conclusion

From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.

     

MIDAS-G: a computational platform for investigating fragmentation rules of tandem mass spectrometry in metabolomics

Introduction

Tandem mass spectrometry (MS/MS) has been widely used for identifying metabolites in many areas. However, computationally identifying metabolites from MS/MS data is challenging due to the unknown of fragmentation rules, which determine the precedence of chemical bond dissociation. Although this problem has been tackled by different ways, the lack of computational tools to flexibly represent adjacent structures of chemical bonds is still a long-term bottleneck for studying fragmentation rules.

Objectives

This study aimed to develop computational methods for investigating fragmentation rules by analyzing annotated MS/MS data.

Methods

We implemented a computational platform, MIDAS-G, for investigating fragmentation rules. MIDAS-G processes a metabolite as a simple graph and uses graph grammars to recognize specific chemical bonds and their adjacent structures. We can apply MIDAS-G to investigate fragmentation rules by adjusting bond weights in the scoring model of the metabolite identification tool and comparing metabolite identification performances.

Results

We used MIDAS-G to investigate four bond types on real annotated MS/MS data in experiments. The experimental results matched data collected from wet labs and literature. The effectiveness of MIDAS-G was confirmed.

Conclusion

We developed a computational platform for investigating fragmentation rules of tandem mass spectrometry. This platform is freely available for download.

     

Chilling slows anaerobic metabolism to improve anoxia tolerance of insects

Background

Insects are renowned for their ability to survive anoxia. Anoxia tolerance may be enhanced during chilling through metabolic suppression.

Aims

Here, the metabolomic response of insects to anoxia, both with and without chilling, for different durations (12–36 h) was examined to assess the potential cross-tolerance mechanisms.

Results

Chilling during anoxia (cold anoxia) significantly improved survival relative to anoxia at warmer temperatures. Reduced intermediate metabolites and increased lactic acid, indicating a switch to anaerobic metabolism, were characteristic of larvae in anoxia.

Conclusions

Anoxia tolerance was correlated survival improvements after cold anoxia were correlated with a reduction in anaerobic metabolism.

     

Biosynthesis of glycerol phosphate is associated with long-term potentiation in hippocampal neurons

Introduction

Neurons have a very high energy requirement, and their metabolism is tightly regulated to ensure delivery of adequate substrate to sustain neuronal activity and neuroplastic changes. The mechanisms underlying the regulation of neuronal metabolism, however, are not completely clear.

Objective

The objective of this study was to investigate the central carbon metabolism in neurons, in order to identify the regulatory pathways governing neuronal anabolism and catabolism.

Methods

Here we first have applied MS-based endometabolomics to elucidate the metabolic dynamics in cultured hippocampal primary neurons. Using nanoLC-ESI-LTQ Orbitrap MS approach followed by statistical analysis, we measure the dynamics of uniformly labeled ¹³C-glucose entering neurons. We adapted the method by coupling offline patch-clamp setup with MS to confirm findings in vivo.

Results

According to non-parametric statistical analysis of metabolic dynamics, in cultured hippocampal neurons, the glycerol phosphate shuttle is active and correlates with the metabolic flux in the pentose phosphate pathway. In the hippocampus, glycerol-3-phosphate biosynthesis was activated in response to long-term potentiation together with the upregulation of glycolysis and the TCA cycle, but was inactive or silenced in basal conditions.

Conclusions

We identified the biosynthesis of glycerol-3-phosphate as a key regulator in mechanisms implicated in learning and memory. Notably, defects in enzymes linked with the glycerol phosphate shuttle have been implicated in neurological disorders and intellectual disability_. These results could improve our understanding of the general mechanisms of learning and memory and facilitate the development of novel therapies for metabolic disorders linked with intellectual disability.

     

A metabolomics approach to uncover effects of different exercise modalities in type 1 diabetes

Introduction

Exercise-associated metabolism in type 1 diabetes (T1D) remains under-studied due to the complex interplay between exogenous insulin, counter-regulatory hormones and insulin-sensitivity.

Objective

To identify the metabolic differences induced by two exercise modalities in T1D using ultra high-performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC–HRMS) based metabolomics.

Methods

Twelve T1D adults performed intermittent high-intensity (IHE) and continuous-moderate-intensity (CONT) exercise. Serum samples were analysed by UHPLC–HRMS.

Results

Metabolic profiling of IHE and CONT highlighted exercise-induced changes in purine and acylcarnitine metabolism.

Conclusion

IHE may increase beta-oxidation through higher ATP-turnover. UHPLC–HRMS based metabolomics as a data-driven approach without an a priori hypothesis may help uncover distinctive metabolic effects during exercise in T1D.Clinical trial registration number is www.clinicaltrials.gov: NCT02068638.

     

Association of mitochondrial dysfunction and lipid metabolism with type 2 diabetes mellitus: A review of literature

Background

Diabetes mellitus (DM) is one of the most prevalent chronic diseases, and its prevalence continues to increase globally. The impact of mitochondrial dysfunction and lipid metabolism on diabetes mellitus and insulin resistance (IR) has been implicated in several previous reports; however, the results of studies are confusing despite four decades of study.

Methods/Results

This review has evaluated updated understanding of the role of mitochondrial dysfunction and lipid metabolism on type 2 diabetes, and found that mitochondrial dysfunction and lipid metabolism disorder induce the dysregulation of liver and pancreatic beta cells, insulin resistance, and type 2 diabetes.

Conclusion

Mitochondrial dysfunction and lipid metabolism induce metabolic dysregulation and finally increasing the possibility of diabetes.

     

Investigation of altered urinary metabolomic profiles of invasive ductal carcinoma of breast using targeted and untargeted approaches

Introduction

Invasive ductal carcinoma (IDC) is a type of breast cancer, usually detected in advanced stages due to its asymptomatic nature which ultimately leads to low survival rate. Identification of urinary metabolic adaptations induced by IDC to understand the disease pathophysiology and monitor therapy response would be a helpful approach in clinical settings. Moreover, its non-invasive and cost effective strategy better suited to minimize apprehension among high risk population.

Objective

This study aims toward investigating the urinary metabolic alterations of IDC by targeted (LC-MRM/MS) and untargeted (GC–MS) approaches for the better understanding of the disease pathophysiology and monitoring therapy response.

Methods

Urinary metabolic alterations of IDC subjects (63) and control subjects (63) were explored by targeted (LC-MRM/MS) and untargeted (GC–MS) approaches. IDC specific urinary metabolomics signature was extracted by applying both univariate and multivariate statistical tools.

Results

Statistical analysis identified 39 urinary metabolites with the highest contribution to metabolomic alterations specific to IDC. Out of which, 19 metabolites were identified from targeted LC-MRM/MS analysis, while 20 were identified from the untargeted GC–MS analysis. Receiver operator characteristic (ROC) curve analysis evidenced 6 most discriminatory metabolites from each type of approach that could differentiate between IDC subjects and controls with higher sensitivity and specificity. Furthermore, metabolic pathway analysis depicted several dysregulated pathways in IDC including sugar, amino acid, nucleotide metabolism, TCA cycle etc.

Conclusions

Overall, this study provides valuable inputs regarding altered urinary metabolites which improved our knowledge on urinary metabolomic alterations induced by IDC. Moreover, this study identified several dysregulated metabolic pathways which offer further insight into the disease pathophysiology.

     

Noninvasive characterization of metabolites secreted in culture media by bovine embryos during in vitro production

Introduction

Secreted molecules could be correlated with the potential of embryonic development. The development of new technologies, such as mass spectrometry (MS), has enabled analyzes in culture medium to favor the determination of embryos viability in order to improve embryo selection.

Objectives

To perform a non-invasive characterization of the secretome of in vitro produced embryos with different kinetics of cleavage and in different stages of development to obtain specific patterns based on embryonic phenotype through MALDI–TOF–MS.

Methods

Bovine embryos were produced in vitro by standard protocols. The zygotes were transferred to individual culture medium and divided into two groups: Fast [4 cells-22 hours past the beginning of culture (hpc)] and Slow (2 cells-22 hpc). Culture media drops were collected at 22, 96 and 168 hpc. Analysis of embryonic secretome was made by MALDI–TOF–MS after extractions of the metabolites. Spectra were acquired in positive ionization mode. Univariate (Fold-change) and multivariate (Partial Least Squares Discriminants Analysis) analyses were performed by the online software Metaboanalyst.

Results

It was demonstrated that embryos with different kinetics have different spectrometric profiles during embryonic development. Moreover, secreted molecules in each developmental stage are differentially represented in embryos with different kinetics, and are related to specific pathways such as lipid and amino acids metabolism and cell proliferation.

Conclusion

We propose that the analysis of culture media by MALDI–TOF–MS can be used for qualitative characterization of bovine embryos, allowing the identification of key molecules during in vitro culture.

     

Optimization of fecal sample preparation for untargeted LC-HRMS based metabolomics

Introduction

Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.

Objectives

In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.

Methods

The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.

Results

A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.

Conclusion

The workflow generated repeatable and informative fingerprints for robust metabolome characterization.

     

Metabolic adaptation following genome doubling in citrus doubled diploids revealed by non-targeted metabolomics

Introduction

Polyploidy is a widespread phenomenon in nature and is thought to play a major role in the evolution of flowering plants. Additionally, polyploidization produces novel phenotypes that through plant breeding have enhanced the production of biomass and improved the stress tolerance of major economic crops. However, the effect of polyploidization on plant metabolism is still unclear.

Objectives

In order to test whether there are common metabolic responses following genome doubling, we performed a comparative metabolomic analysis of mature leaves from doubled diploids and the corresponding diploids of red tangerine (Citrus reticulata Blanco), trifoliate orange (Poncirus trifoliata L. Raf.) and precocious trifoliate orange (P. trifoliata).

Methods

Non-targeted and targeted metabolic profiling of mature leaves from three doubled diploids and their diploid controls were performed by using liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS) and gas chromatography–mass spectrometry (GC–MS).

Results

About 11–34% of the detected metabolic features differentially accumulated in the doubled diploids, mostly by less than fivefold. The levels of primary metabolites tended to increase in the doubled diploids. Concentrations of tricarboxylic acid cycle intermediates—citric acid, malic acid, fumaric acid and succinic acid, enhanced in all of the doubled diploids. The levels of secondary metabolites, including phenylpropanoids and terpenoids, tended to decrease in the doubled diploids. This is consistent with the lower C/N ratios in the doubled diploids.

Conclusions

Polyploidization had a significant but relatively limited influence on the accumulation of metabolites in these citrus species. We conclude that primary metabolism takes priority over secondary metabolism in doubled diploid plants to relieve the “genomic stress” encountered during the early stages of genome doubling, probably to promote vitality and growth.

     

The Turkish validation of the Brief International Cognitive Assessment for Multiple Sclerosis (BICAMS) battery

Background

Cognitive impairment may be seen in as many as 43–70% of patients with multiple sclerosis (MS) and may be observed in all MS subtypes. The Brief International Cognitive Assessment in Multiple Sclerosis (BICAMS) battery may be used to evaluate cognition status. The purpose of the current study is to validate the BICAMS battery in Turkish.

Methods

Patients with MS attending our clinic between September 2014 and April 2015 were invited to participate. Healthy control participants were matched in terms of age, gender and years of education.

Results

One hundred seventy-three MS patients and 153 healthy control participants were enrolled in the study. MS patients performed significantly worse in all trials than the members of the healthy control group. In addition, cognitive dysfunction was identified in 78 of the 173 (45.1%) patients. In the MS with cognitive impairment group, 64 out of 151 (42.4%) subjects were RRMS patients, 12 out of 18 (66.7%) were secondary progressive MS patients, and 2 out of 4 (50%) were primer progressive MS patients.

Conclusions

The BICAMS has been proposed for assessing cognitive impairment in MS patients. This study shows that the battery is suitable for use in Turkey.

     

GC–MS analysis of the ruminal metabolome response to thiamine supplementation during high grain feeding in dairy cows

Introduction

Thiamine is known to attenuate high-concentrate diet induced subacute ruminal acidosis (SARA) in dairy cows, however, the underlying mechanisms remain unclear.

Objectives

The major objective of this study was to investigate the metabolic mechanisms of thiamine supplementation on high-concentrate diet induced SARA.

Methods

Six multiparous, rumen-fistulated Holstein cows were used in a replicated 3?×?3 Latin square design. The treatments included a control diet (CON; 20% starch, dry matter basis), a SARA-inducing diet (SAID; 33.2% starch, dry matter basis) and SARA-inducing diet supplemented with 180 mg of thiamine/kg of dry matter intake (SAID?+?T). On d21 of each period, ruminal fluid samples were collected at 3 h post feeding, and GC/MS was used to analyze rumen fluid samples.

Results

PCA and OPLS-DA analysis demonstrated that the ruminal metabolite profile were different in three treatments. Compared with CON treatment, SAID feeding significantly decreased rumen pH, acetate, succinic acid, increased propionate, pyruvate, lactate, glycine and biogenic amines including spermidine and putrescine. Thiamine supplementation significantly decreased rumen content of propionate, pyruvate, lactate, glycine and spermidine; increase rumen pH, acetate and some medium-chain fatty acids. The enrichment analysis of different metabolites indicated that thiamine supplementation mainly affected carbohydrates, amino acids, pyruvate and thiamine metabolism compared with SAID treatment.

Conclusions

These findings revealed that thiamine supplementation could attenuate high-concentrate diet induced SARA by increasing pyruvate formate-lyase activity to promote pyruvate to generate acetyl-CoA and inhibit lactate generation. Besides, thiamine reduced biogenic amines to alleviate ruminal epithelial inflammatory response.

     

Metabolomics of biomarker discovery in ovarian cancer: a systematic review of the current literature

Introduction

Metabolomics is the emerging member of “omics” sciences advancing the understanding, diagnosis and treatment of many cancers, including ovarian cancer (OC).

Objectives

To systematically identify the metabolomic abnormalities in OC detection, and the dominant metabolic pathways associated with the observed alterations.

Methods

An electronic literature search was performed, up to and including January 15th 2016, for studies evaluating the metabolomic profile of patients with OC compared to controls. QUADOMICS tool was used to assess the quality of the twenty-three studies included in this systematic review.

Results

Biological samples utilized for metabolomic analysis include: serum/plasma (n = 13), urine (n = 4), cyst fluid (n = 3), tissue (n = 2) and ascitic fluid (n = 1). Metabolites related to cellular respiration, carbohydrate, lipid, protein and nucleotide metabolism were significantly altered in OC. Increased levels of tricarboxylic acid cycle intermediates and altered metabolites of the glycolytic pathway pointed to perturbations in cellular respiration. Alterations in lipid metabolism included enhanced fatty acid oxidation, abnormal levels of glycerolipids, sphingolipids and free fatty acids with common elevations of palmitate, oleate, and myristate. Increased levels of glutamine, glycine, cysteine and threonine were commonly reported while enhanced degradations of tryptophan, histidine and phenylalanine were found. N-acetylaspartate, a brain amino acid, was found elevated in primary and metastatic OC tissue and ovarian cyst fluid. Further, elevated levels of ketone bodies including 3-hydroxybutyrate were commonly reported. Increased levels of nucleotide metabolites and tocopherols were consistent through out the studies.

Conclusion

Metabolomics presents significant new opportunities for diagnostic biomarker development, elucidating previously unknown mechanisms of OC pathogenesis.

     

Polycystic ovary syndrome in Indian women: a mass spectrometry based serum metabolomics approach

Introduction

Polycystic ovary syndrome (PCOS) is a complex, heterogeneous endocrinological disorder with uncertain pathogenesis and is very common in women of reproductive age. There are few reports of utilizing metabolomics approach to understand the complex pathophysiology of PCOS. However, excluding one previous NMR-based metabolomics study, none of the study was conducted in Indian population.

Objective

The study aims to compare the serum metabolomic profile of PCOS women with controls from the Eastern region of India.

Methods

PCOS women (n?=?35) and healthy control women (n?=?30) undergoing tubal ligation were recruited for this study. Serum metabolic profiles were generated using liquid chromatography–tandem mass spectrometry (LC-MS/MS) and gas chromatography–mass spectrometry (GC-MS). Multivariate statistical analysis was applied to spectral data obtained from both the LC-MS/MS and GC/MS.

Results

Nine metabolites were identified to be most significantly dysregulated in sera of PCOS women; however, few other identified metabolites were also altered but with lesser significance. Amongst these metabolites, riboflavin, sucrose, adenine and N-acetyl glycine, phosphoric acid and cortisol were down-regulated, whereas, thymine, cystathionine, and phenylalanine were up-regulated in PCOS when compared with controls. The observed changes in metabolite expression suggested alterations in aminoacyl-tRNA biosynthesis, metabolism of nitrogen, alanine-aspartate-glutamate, galactose, glycine-serine-threonine, and pyrimidine-purine among several metabolic pathways possibly implicated in these PCOS women.

Conclusion

The altered metabolites identified in PCOS women of Eastern Indian population, provide insight into current perceptive of the disease pathology, metabolic involvements, and may be considered as putative markers of PCOS.

     

The comparison of insulin and uric acid levels in adolescents with and without metabolic syndrome

Background and Aim

The prevalence of metabolic syndrome (MS) increased in recent years in both adolescents and children groups. The aim of the study is evaluating the relationship between insulin and uric acid (UA) level in MS in adolescents

Materials and Methods

we studied 120 adolescence aged 10 to 19 in two groups: control group without metabolic syndrome and case group with metabolic syndrome. The Criteria of ATP III was considered as a diagnosis factor for metabolic syndrome.

Discussion

Various studies have been conducted in various populations to evaluate the relationship between UA level and MS in adolescents. Abdominal obesity, low HDL, hypertriglyceridemia and hypertension are associated with high UA level. In their analysis, the MS OR in UA level?4.9, 4.9-5.8 and ?5.8 mg/dl was 1, 2.53 and 9.03, respectively, which were higher than our findings in current study. Hyperinsulinemia caused by insulin resistance is one of the complications associated with MS, which puts individuals at risk of diabetes and cardiovascular events.

Results

Uric acid level in the Case group was significantly higher than the control group (p = 0.0001, 43.8±1.4 vs. 4.1±1 mg/dl, respectively). Insulin level was significantly higher in the case group in compare to the control group (p = 0.008, 9.8± 5.3 vs. 12.2±6 μU/ml, respectively).

Conclusion

The findings of this case-control study showed that adolescents with metabolic syndrome have a higher uric acid and insulin level in compare to normal subjects. We hypothesis that increase in serum insulin and uric acid level can be a risk factor in the development of metabolic syndrome.

     

Crystal structure of <Emphasis Type="Italic">Escherichia coli</Emphasis> protein ybgI,a toroidal structure with a dinuclear metal site

Background

The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results

The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion

The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

     

Circulating sterols as predictors of early allograft dysfunction and clinical outcome in patients undergoing liver transplantation

Introduction

Sensitive and specific assessment of the hepatic graft metabolism after liver transplantation (LTX) is essential for early detection of postoperative dysfunction implying the need for consecutive therapeutic interventions.

Objectives

Here, we assessed circulating liver metabolites of the cholesterol pathway, amino acids and acylcarnitines and evaluated their predictive value on early allograft dysfunction (EAD) and clinical outcome in the context of LTX.

Methods

The metabolites were quantified in the plasma of 40 liver graft recipients one day pre- and 10 days post-LTX by liquid chromatography/tandem mass spectrometry (LC–MS/MS). Plant sterols as well as cholesterol and its precursors were determined in the free and esterified form; lanosterol in the free form only. Metabolites and esterification ratios were compared to the model for early allograft function scoring (MEAF) which is calculated at day 3 post-LTX from routine parameters defining EAD.

Results

The hepatic esterification ratio of all sterols, but not amino acids and acylcarnitine concentrations, showed substantial metabolic disturbances post-LTX and correlated to the MEAF. In ROC analysis, the low esterification ratio of β-sitosterol and stigmasterol from day 1 and of the other sterols from day 3 were predictive for a high MEAF, i.e. EAD. Additionally, the ratio of esterified β-sitosterol and free lanosterol were predictive for all days and the esterification ratio of the other sterols at day 3 or 4 post-LTX for 3-month mortality.

Conclusion

Low ratios of circulating esterified sterols are associated with a high risk of EAD and impaired clinical outcome in the early postoperative phase following LTX.