High-Throughput RPLC-MS/MS Quantification of Short- and Medium-Chain Fatty Acids.

Short- and medium-chain fatty acids (SMCFA) are monocarboxylic acids with a carbon chain length of 1-12 carbon atoms. They are mainly produced in humans by the gut microbiota, play crucial metabolic roles, are vital for intestinal health, and have multifaceted impact on immune and neurological functions. Accurate detection and quantification of SMCFA in different human biofluids is achieved using 3-nitro phenylhydrazine (3-NPH) derivatization of the free fatty acids followed by reverse phase liquid chromatography (RPLC) separation and detection by tandem mass spectrometry (MS/MS). Here, we describe the simultaneous measurement of 14 SMCFA and lactate in detail. All 3-NPH-SMCFA-hydrazones are separated in less than 5 min with an 8-min total run time (injection-to-injection). Linear dynamic range over 0.1-500 µM is achieved for most SCFAs, while it is 0.05-100 µM for MCFAs. Validation of the procedure depicts good linearity (R2 > 0.98) and repeatability (CV ≤ 20%). The lower limit of detection (LLOD) is 10-30 nM. The lower limit of quantification (LLOQ) is 50-100 nM for most analytes, while it is 0.5 µM for acetate. In conclusion, the method offers several benefits compared to alternative methods regarding throughput, selectivity, sensitivity, and robustness.

Rapid Analysis of Sterols in Blood-Derived Samples.

Dysregulations of cholesterol biosynthesis are known to be associated with several pathologies. Due to the rapid growth of clinical investigations in this research area, a specific, fast, and valid method for analyzing cholesterol, its precursors, and metabolites is required. Here, we describe a rapid method for sample preparation, separation, and quantification of sterols in blood-derived samples using polymeric solid phase extraction followed by gas chromatography-mass spectrometry. The validated method demonstrates a reliable quantification of cholesterol, its precursors, and metabolites.

Uncovering the mechanisms of Zhubi decoction against rheumatoid arthritis through an integrated study of network pharmacology, metabolomics, and intestinal flora.

ETHNOPHARMACOLOGICAL RELEVANCE: Zhubi Decoction (ZBD) is a modified formulation derived from the classic traditional Chinese medicine prescription "Er-Xian Decoction" documented in the esteemed "Clinical Manual of Chinese Medical Prescription". While the utilization of ZBD has exhibited promising clinical outcomes in treating rheumatoid arthritis (RA), the precise bioactive chemical constituents and the underlying mechanisms involved in its therapeutic efficacy remain to be comprehensively determined. AIM OF THE STUDY: This study aims to systematically examine ZBD's pharmacological effects and molecular mechanisms for RA alleviation. MATERIALS AND METHODS: Utilizing the collagen-induced arthritis (CIA) rat model, we comprehensively evaluated the anti-rheumatoid arthritis effects of ZBD in vivo through various indices, such as paw edema, arthritis index, ankle diameter, inflammatory cytokine levels, pathological conditions, and micro-CT analysis. The UPLC-MS/MS technique was utilized to analyze the compounds of ZBD. The potential therapeutic targets and signaling pathways of ZBD in the management of RA were predicted using network pharmacology. To analyze comprehensive metabolic profiles and identify underlying metabolic pathways, we conducted a serum-based widely targeted metabolomics analysis utilizing LC-MS technology. Key targets and predicted pathways were further validated using immunofluorescent staining, which integrated findings from serum metabolomics and network pharmacology analysis. Additionally, we analyzed the gut microbiota composition in rats employing 16 S rDNA sequencing and investigated the effects of ZBD on the microbiota of CIA rats through bioinformatics and statistical methods. RESULTS: ZBD exhibited remarkable efficacy in alleviating RA symptoms in CIA rats without notable side effects. This included reduced paw redness and swelling, minimized joint damage, improved the histopathology of cartilage and synovium, mitigated the inflammatory state, and lowered serum concentrations of cytokines TNF-α, IL-1ß and IL-6. Notably, the effectiveness of ZBD was comparable to MTX. Network pharmacology analysis revealed inflammation and immunity-related signaling pathways, such as PI3K/AKT, MAPK, IL-17, and TNF signaling pathways, as vital mediators in the effectual mechanisms of ZBD. Immunofluorescence analysis validated ZBD's ability to inhibit PI3K/AKT pathway proteins. Serum metabolomics studies revealed that ZBD modulates 170 differential metabolites, partially restored disrupted metabolic profiles in CIA rats. With a notable impact on amino acids and their metabolites, and lipids and lipid-like molecules. Integrated analysis of metabolomics and network pharmacology identified 6 pivotal metabolite pathways and 3 crucial targets: PTGS2, GSTP1, and ALDH2. Additionally, 16 S rDNA sequencing illuminated that ZBD mitigated gut microbiota dysbiosis in the CIA group, highlighting key genera such as Ligilactobacillus, Prevotella_9, unclassified_Bacilli, and unclassified_rumen_bacterium_JW32. Correlation analysis disclosed a significant link between 47 distinct metabolites and specific bacterial species. CONCLUSION: ZBD is a safe and efficacious TCM formulation, demonstrates efficacy in treating RA through its multi-component, multi-target, and multi-pathway mechanisms. The regulation of inflammation and immunity-related signaling pathways constitutes a crucial mechanism of ZBD's efficacy. Furthermore, ZBD modulates host metabolism and intestinal flora. The integrated analysis presents experimental evidence of ZBD for the management of RA.

Evaluation and the mechanism of ShengXian and JinShuiLiuJun decoction in the treatment of silicotic fibrosis: An integrated network pharmacology, life omics, and experimental validation study.

BACKGROUND: Silicosis is a systemic disease characterized by extensive fibrosis due to prolonged exposure to silica dust, with rising incidence rates significantly impacting global public health. ShengXian and JinShuiLiuJun Decoction (SXD) is a Chinese medicinal preparation containing a variety of medicinal plants. It has shown notable clinical efficacy in treating silicotic fibrosis in China. However, the precise mechanisms underlying its therapeutic effects remain unclear. This study integrates network pharmacology, multi-omics analysis, and experimental validation to investigate the potential mechanisms by which SXD treats silicotic fibrosis. OBJECTIVE: The study aims to investigate the therapeutic efficacy of SXD in treating silicotic fibrosis and to elucidate its underlying molecular mechanisms. METHODS: HPLC-Q-TOF-MS was used to identify the active components of SXD, and combined with network pharmacology, metabolomics, and transcriptomics, the mechanism of SXD in treating silicotic fibrosis was explored from multiple perspectives. The therapeutic effect of SXD was assessed through HE staining, Masson staining, Micro CT imaging, pulmonary function tests, and hydroxyproline content in lung tissue. Finally, network pharmacology and multi-omics findings were validated using molecular docking. CETSA, immunofluorescence, SPR, and Western blotting were used to analyze key factors in the NF-κB pathway at the animal, cellular, and molecular levels. RESULTS: SXD treatment improved lung function in silicosis rats, reduced inflammatory cell infiltration, collagen deposition, fibrosis and other pathological changes, and inhibited the protein expression of TNF-α, IL-17A, and IL-1ß, and NF-κB in lung tissue. HPLC-Q-TOF-MS combined with network pharmacology identified key compounds such as Liquiritigenin, 3-Methoxynobiletin, Isomangiferin, Hesperidin, shogaol, and Ligustroflavone, which likely exert therapeutic effects through the TNF, IL-17, NF-κB, and TGF-ß signaling pathways. Transcriptomics and metabolomics results revealed that SXD up-regulated the expression of NF-κB pathway-related genes (NFKBIA, NFKBIZ) and key regulators of the retinol metabolism pathway, while down-regulating pro-inflammatory genes (IL1B, IL17A, IL6). Experimental findings confirmed that SXD suppressed the expression of NF-κB pathway-related proteins and upstream activators TNF-α, IL-17A, and IL-1ß, as well as their receptors, in both lung tissue and cellular models. Additionally, SXD-containing serum had a direct, non-toxic effect on MRC-5 cells, effectively inhibiting collagen expression and TGF-ß secretion. SXD also had a positive effect on collagen production and extracellular matrix (ECM) aggregation in fibroblasts. Molecular dynamics studies showed that SXD directly binds to NF-κB and IκB. CONCLUSION: SXD exerts therapeutic effects on silicotic fibrosis by inhibiting NF-κB signaling transduction mediated by TNF-α, IL-17A, and IL-1ß, and suppressing fibroblast activation.

Serum metabolite biomarkers for the early diagnosis and monitoring of age-related macular degeneration.

INTRODUCTION: Age-related macular degeneration (AMD) is a leading cause of irreversible blindness worldwide, with significant challenges for early diagnosis and treatment. OBJECTIVES: To identify new biomarkers that are important for the early diagnosis and monitoring of the severity/progression of AMD. METHODS: We investigated the diagnostic and monitoring potential of blood metabolites in a cohort of 547 individuals (167 healthy controls, 240 individuals with other eye diseases as eye disease controls, and 140 individuals with AMD) from 2 centers over three phases: discovery phase 1, discovery phase 2, and an external validation phase. The samples were analyzed via a mass spectrometry-based, widely targeted metabolomic workflow. In discovery phases 1 and 2, we built a machine learning algorithm to predict the probability of AMD. In the external validation phase, we further confirmed the performance of the biomarker panel identified by the algorithm. We subsequently evaluated the performance of the identified biomarker panel in monitoring the progression and severity of AMD. RESULTS: We developed a clinically specific three-metabolite panel (hypoxanthine, 2-furoylglycine, and 1-hexadecyl-2-azelaoyl-sn-glycero-3-phosphocholine) via five machine learning models. The random forest model effectively discriminated patients with AMD from patents in the other two groups and showed acceptable calibration (area under the curve (AUC) = 1.0; accuracy = 1.0) in both discovery phases 1 and 2. An independent validation phase confirmed the diagnostic model's efficacy (AUC = 0.962; accuracy = 0.88). The three-biomarker panel model demonstrated an AUC of 1.0 in differentiating the severity of AMD via RF machine learning, which was consistent across both the discovery and external validation phases. Additionally, the biomarker concentrations remained stable under repeated freeze-thaw cycles (P > 0.05). CONCLUSIONS: This study reveals distinct metabolite variations in the serum of AMD patients, paving the way for the development of the first routine laboratory test for AMD.

Glutamine Attenuates Inflammation and Stimulates Amniotic Cell Proliferation in Premature Rupture of Membranes-related in vitro Models.

Premature rupture of membranes (PROM), with a prevalence of 15.3% in China, frequently results in adverse pregnancy outcomes. In this study, we aimed to identify amino acid metabolites that were differentially expressed in PROM versus healthy controls (HC) using targeted metabolomics and further explored their mechanisms of action with in vitro models.Inclusion and exclusion criteria were established to recruit 50 PROM and 50 HC cases for targeted metabolomics analysis. Twenty-three amino acid metabolites were quantified in the secretions of the posterior vaginal fornix of pregnant women between 31 and 36 weeks of gestation. Glutamine (0.0216 vs. 0.037 µg/mg, P = 0.003, AUC = 72.1%) was identified as the most differentially expressed amino acid metabolite between PROM and HC groups, and had a negative correlation with the abundance of Gardnerella (r=-0.3868, P = 0.0055), Megasphaera (r=-0.3130, P = 0.0269), and Prevotella (r=-0.2944, P = 0.0380), respectively.In amniotic epithelial cell and macrophage co-culture model, Glutamine reduced inflammatory cytokines and chemokines expression and suppressed macrophage chemotaxis. In LPS stimulated RAW 264.7 inflammation model, Glutamine inhibited the expression of inflammatory proteins iNOS and COX-2, down-regulated mRNA transcription of TNF, IL-6, and IL-1ß, and reduced the production of reactive oxygen species through inhibiting NF-κB signaling pathway, and therefore demonstrated its anti-inflammatory effect. Furthermore, Glutamine protected amniotic epithelial cell from autophagy and stimulated its proliferation, therefore may intensify fetal membrane and prevent PROM in vivo.Our results suggested that low Glutamine level in vaginal secretion can be used as an indicator for PROM, and local Glutamine supplementation is a potential intervention and prevention strategy for PROM.

Infants with biliary atresia exhibit an altered amino acid profile in their newborn screening.

INTRODUCTION: Biliary atresia (BA) is a rare progressive neonatal cholangiopathy with unknown pathophysiology and time of onset. Newborn Screening (NBS) in Germany is routinely performed in the first days of life to identify rare congenital diseases utilizing dried blood spot (DBS) card analyses. Infants with biliary atresia (BA) are known to have altered amino acid profiles (AAP) at the time point of diagnosis, but it is unclear whether these alterations are present at the time point of NBS. OBJECTIVES: We aimed to analyze amino acid profiles in NBS-DBS of infants with Biliary Atresia. METHODS: Original NBS-DBS cards of 41 infants who were later on diagnosed with BA were retrospectively obtained. NBS-DBS cards from healthy newborns (n = 40) served as controls. In some BA infants (n = 14) a second DBS card was obtained at time of Kasai surgery. AAP in DBS cards were analyzed by targeted metabolomics. RESULTS: DBS metabolomics in the NBS of at that time point seemingly healthy infants later diagnosed with BA revealed significantly higher levels of Methionine (14.6 ± 8.6 µmol/l), Histidine (23.5 ± 50.3 µmol/l), Threonine (123.9 ± 72.8 µmol/l) and Arginine (14.1 ± 11.8 µmol/l) compared to healthy controls (Met: 8.1 ± 2.6 µmol/l, His: 18.6 ± 10.1 µmol/l, Thr: 98.1 ± 34.3 µmol/l, Arg: 9.3 ± 6.6 µmol/l). Methionine, Arginine and Histidine showed a further increase at time point of Kasai procedure. No correlation between amino acid levels and clinical course was observed. CONCLUSION: Our data demonstrate that BA patients exhibit an altered AAP within 72 h after birth, long before the infants become symptomatic. This supports the theory of a prenatal onset of the disease and, thus, the possibility of developing a sensitive and specific NBS. Methionine might be particularly relevant due to its involvement in glutathione metabolism. Further investigation of AAP in BA may help in understanding the underlying pathophysiology.

Comparison of the difference in the anti-inflammatory activity of two different color types of Farfarae Flos based on in vitro, in vivo experiments and untargeted metabolomics.

Introduction: Due to its remarkable anti-inflammatory pharmacological activity, Farfarae Flos has gained extensive usage in the treatment of various inflammatory diseases such as bronchitis, pneumonia, prostatitis and colitis. And Farfarae Flos come in two color types depending on the color of the flowers: yellowish-white (YW), and purplish-red (PR). However, the difference in anti-inflammatory activity and metabolic profiles between the two flower colors remains unexplored. Methods: This study aims to explore the difference in the anti-inflammatory potential between YW and PR variants of Farfarae Flos and unravel the mechanisms responsible for the observed differences in anti-inflammatory activity through an integrated approach encompassing untargeted metabolomics and in vivo/vitro experimental studies. Initially, we verified the contrasting effects of YW and PR on the inhibition of the inflammatory factors interleukin-6 (IL-6) and nitric oxide (NO) by utilizing an in vitro RAW 264.7 cell inflammation model. Subsequently, a comprehensive evaluation of the systemic inhibitory capacity of YW and PR on IL-6, Interleukin-10 (IL-10), and tumor necrosis factor-α (TNF-α) was conducted using a validated whole-body mouse model, followed by the analysis of inflammatory factors and histological examination of collected serum, liver, and spleen after 7 days. Furthermore, non-targeted metabolomics profiling was employed to analyze the metabolite profiles of Farfarae Flos with different colors, and quantitative analysis was conducted to identify differential metabolites between YW and PR. The correlation between the anti-inflammatory activities of differentially accumulated metabolites (DAMs) and Farfarae Flos was investigated, resulting in the identification of 48 compounds exhibiting significant anti-inflammatory activity. Additionally, KEGG pathway enrichment analysis was performed to elucidate the underlying mechanisms. Results: Our findings demonstrate that both YW and PR possess anti-inflammatory abilities, with PR exhibiting significantly superior efficacy. The integration of in vivo/vitro experiments and non-targeted metabolomics confirmed the exceptional anti-inflammatory potential of PR and solidified its classification as the "purplish-red better" of Farfarae Flos. Discussion: This study provides valuable insights into the breeding and medical transformation of Farfarae Flos varieties, along with a scientific basis for the establishment of quality standards and the development of new drugs utilizing Farfarae Flos.

Widely Targeted Metabolomic Analysis Reveals Dynamic Metabolic Changes in Yanbian Cattle during Dry-Aging Process.

Dry-aging is a postmortem process that can substantially enhance the texture and flavour of beef. This study entailed suspending Yanbian cattle M. gluteus medius in the aging cabinet, maintained at a temperature of 2-4 °C and a relative humidity of 85 ± 5% for 35 days. Throughout this period, samples were systematically collected every 7 days. The widely targeted metabolomic analysis has been used in this investigation to analyse the dynamic changes in Yanbian cattle metabolites during dry-aging. A total of 883 metabolites were identified, with amino acids and their metabolites representing the largest proportion. Multivariate statistical analysis showed that 373 metabolites were identified as differential metabolites that changed significantly during the dry-aging process, including metabolites of amino acids, glycerophospholipids, and nucleotides and their metabolites. Additionally, 308 metabolites exhibited various increasing trends with time in dry-aging. The analysis of KEGG pathway analysis showed that ABC transporters, glycerophospholipid, and arachidonic acid metabolism are the most important metabolic pathways during dry-aging. These findings can guide technological developments in the meat processing sector and provide valuable insights into the metabolic traits and pathways of Yanbian cattle during the dry-aging process.

Changes in Quality Characteristics and Metabolite Composition of Low-Temperature and Nitrogen-Modified Atmosphere in Indica Rice during Storage.

A low temperature (LT) is used to delay grain deterioration effectively. In practical applications, a nitrogen-modified atmosphere (N2) is also an effective way of preventing grain pests and delaying grain deterioration. However, there are few studies on grain quality changes using a combination treatment of an LT and N2 during storage. In this study, the storage quality, processing characteristics, and metabolites of rice under conventional storage (CS), LT (20 °C), N2 (95%), and LT+N2 treatments were analyzed for 180 days, under a controlled humidity of 65% ± 2%. The results showed that compared to the CS, LT, and N2 treatments, the LT+N2 treatment had the best effect in retarding the increase in MDA and electrical conductivity and deferring the decrease in CAT activity. In addition, the LT+N2 treatment maintained the color of the rice better and sustained a better processing quality. Non-targeted metabolomics analysis further confirmed that the LT+N2 treatment maintained the vigor of the rice and retarded its spoilage by activating the metabolisms of amino acids, carbohydrates, and flavonoids. These results suggest a favorable practice for preventing storage deterioration and increasing the processing quality for rice storage. They provided new insights into the mechanisms of rice quality changes using the combination treatment of an LT and N2.

A simplified metabolomic analysis of dried blood spots in breast cancer patients.

Breast cancer (BC) is among the most commonly diagnosed cancers. Besides mammography, breast ultrasonography and the routinely monitored protein markers, the variations of small molecular metabolites in blood may be of great diagnostic value. This study aimed to quantify specific metabolite markers with potential application in BC detection. The study enrolled 50 participants, 25 BC patients and 25 healthy controls (CTRL). Dried blood spots (DBS) were utilized as biological media and were quantified via a simplified liquid chromatography tandem mass spectrometry (LC-MS/MS) method, used in expanded newborn screening. The targeted metabolomic analysis included 12 amino acids and 32 acylcarnitines. Statistical analysis revealed a significant variation of metabolic profiles between BC patients and CTRL. Among the 44 metabolites, 18 acylcarnitines and 10 amino acids remained significant after Bonferroni correction, showing increase or decrease and enabled classification of BC patients and CTRL. The well-established LC-MS/MS protocol could provide results within few minutes. Therefore, the combination of an easy-to-handle material-DBS and LC-MS/MS protocol could facilitate BC screening/diagnosis and in the next step applied to other cancer patients, as well.

Sulfated Bile Acids in Serum as Potential Biomarkers of Disease Severity and Mortality in COVID-19.

The fight against coronavirus disease 2019 (COVID-19) continues. Since the pandemic's onset, several biomarkers have been proposed to assess the diagnosis and prognosis of this disease. This research aimed to identify potential disease severity biomarkers in serum samples of patients with COVID-19 during the disease course. Data were collected using untargeted and targeted mass spectrometry methods. The results were interpreted by performing univariate and multivariate analyses. Important metabolite classes were identified by qualitative untargeted metabolomics in 15 serum samples from survivors of COVID-19. Quantitative targeted metabolomics on a larger patient cohort including 15 non-survivors confirmed serum 3-sulfate bile acids (i.e. GLCA-3S) were significantly increased in non-survivors compared to survivors during the early disease stage (p-value < 0.0001). Notably, it was associated with a higher risk of mortality (odds ratio of 26). A principal component analysis showed the ability to discriminate between survivors and non-survivors using the BA concentrations. Furthermore, increased BA-S is highly correlated with known parameters altered in severe clinical conditions.

Exploring the Diversity and Specificity of Secondary Biosynthetic Potential in Rhodococcus.

The actinomycete genus Rhodococcus is known for its diverse biosynthetic enzymes, with potential in pollutant degradation, chemical biocatalysis, and natural product exploration. Comparative genomics have analyzed the distribution patterns of non-ribosomal peptide synthetases (NRPSs) in Rhodococcus. The diversity and specificity of its secondary metabolism offer valuable insights for exploring natural products, yet remain understudied. In the present study, we analyzed the distribution patterns of biosynthetic gene clusters (BGCs) in the most comprehensive Rhodococcus genome data to date. The results show that 86.5% of the gene cluster families (GCFs) are only distributed in a specific phylogenomic-clade of Rhodococcus, with the most predominant types of gene clusters being NRPS and ribosomally synthesized and post-translationally modified peptides (RiPPs). In-depth mining of RiPP gene clusters revealed that Rhodococcus encodes many clade-specific novel RiPPs, with thirteen core peptides showing antibacterial potential. High-throughput elicitor screening (HiTES) and non-targeted metabolomics revealed that a marine-derived Rhodococcus strain produces a large number of new aurachin-like compounds when exposed to specific elicitors. The present study highlights the diversity and specificity of secondary biosynthetic potential in Rhodococcus, and provides valuable information for the targeted exploration of novel natural products from Rhodococcus, especially for phylogenomic-clade-specific metabolites.

Role of human plasma metabolites in prediabetes and type 2 diabetes from the IMI-DIRECT study.

AIMS/HYPOTHESIS: Type 2 diabetes is a chronic condition that is caused by hyperglycaemia. Our aim was to characterise the metabolomics to find their association with the glycaemic spectrum and find a causal relationship between metabolites and type 2 diabetes. METHODS: As part of the Innovative Medicines Initiative - Diabetes Research on Patient Stratification (IMI-DIRECT) consortium, 3000 plasma samples were measured with the Biocrates AbsoluteIDQ p150 Kit and Metabolon analytics. A total of 911 metabolites (132 targeted metabolomics, 779 untargeted metabolomics) passed the quality control. Multivariable linear and logistic regression analysis estimates were calculated from the concentration/peak areas of each metabolite as an explanatory variable and the glycaemic status as a dependent variable. This analysis was adjusted for age, sex, BMI, study centre in the basic model, and additionally for alcohol, smoking, BP, fasting HDL-cholesterol and fasting triacylglycerol in the full model. Statistical significance was Bonferroni corrected throughout. Beyond associations, we investigated the mediation effect and causal effects for which causal mediation test and two-sample Mendelian randomisation (2SMR) methods were used, respectively. RESULTS: In the targeted metabolomics, we observed four (15), 34 (99) and 50 (108) metabolites (number of metabolites observed in untargeted metabolomics appear in parentheses) that were significantly different when comparing normal glucose regulation vs impaired glucose regulation/prediabetes, normal glucose regulation vs type 2 diabetes, and impaired glucose regulation vs type 2 diabetes, respectively. Significant metabolites were mainly branched-chain amino acids (BCAAs), with some derivatised BCAAs, lipids, xenobiotics and a few unknowns. Metabolites such as lysophosphatidylcholine a C17:0, sum of hexoses, amino acids from BCAA metabolism (including leucine, isoleucine, valine, N-lactoylvaline, N-lactoylleucine and formiminoglutamate) and lactate, as well as an unknown metabolite (X-24295), were associated with HbA1c progression rate and were significant mediators of type 2 diabetes from baseline to 18 and 48 months of follow-up. 2SMR was used to estimate the causal effect of an exposure on an outcome using summary statistics from UK Biobank genome-wide association studies. We found that type 2 diabetes had a causal effect on the levels of three metabolites (hexose, glutamate and caproate [fatty acid (FA) 6:0]), whereas lipids such as specific phosphatidylcholines (PCs) (namely PC aa C36:2, PC aa C36:5, PC ae C36:3 and PC ae C34:3) as well as the two n-3 fatty acids stearidonate (18:4n3) and docosapentaenoate (22:5n3) potentially had a causal role in the development of type 2 diabetes. CONCLUSIONS/INTERPRETATION: Our findings identify known BCAAs and lipids, along with novel N-lactoyl-amino acid metabolites, significantly associated with prediabetes and diabetes, that mediate the effect of diabetes from baseline to follow-up (18 and 48 months). Causal inference using genetic variants shows the role of lipid metabolism and n-3 fatty acids as being causal for metabolite-to-type 2 diabetes whereas the sum of hexoses is causal for type 2 diabetes-to-metabolite. Identified metabolite markers are useful for stratifying individuals based on their risk progression and should enable targeted interventions.

Bioavailability and interactions of schisandrin B with 5-fluorouracil in a xenograft mouse model of colorectal cancer.

Schisandrin B (Sch B) is a predominant bioactive lignan from the fruit of a Chinese medicine food homology plant, Schisandra chinensis. Previously, we observed potent anti-tumor effect of Sch-B in colorectal cancer (CRC) and enhanced chemotherapy efficacy with fluorouracil (5-FU). However, their bioavailability and reciprocal interactions under CRC conditions are unclear. In this study, we first compared the bioavailability, metabolism and tissue distribution of Sch-B between non-tumor-bearing and xenograft CRC tumor-bearing mice. Next, we examined SchB-5-FU interactions via investigating alterations in drug metabolism and multidrug resistance. Using a validated targeted metabolomics approach, five active metabolites, including Sch-B and fluorodeoxyuridine triphosphate, were found tumor-accumulative. Co-treatment resulted in higher levels of Sch-B and 5-FU metabolites, showing improved phytochemical and drug bioavailability. Multidrug resistance gene (MDR1) was significantly downregulated upon co-treatment. Overall, we demonstrated the potential of Sch-B to serve as a promising chemotherapy adjuvant via improving drug bioavailability and metabolism, and attenuating MDR.

Longitudinal characterization of the muscle metabolome in dairy cows during the transition from lactation cessation to lactation resumption.

Skeletal muscle is vital in maintaining metabolic homeostasis and adapting to the physiological needs of pregnancy and lactation. Despite advancements in understanding metabolic changes in dairy cows around calving and early lactation, there are still gaps in our knowledge, especially concerning muscle metabolism and the changes associated with drying off. This study aimed to characterize the skeletal muscle metabolome in the context of the dietary and metabolic changes occurring during the transition from the cessation of lactation to the resumption of lactation in dairy cows. Twelve Holstein dairy cows housed in tie stalls were dried off 6 weeks (wk) before the expected calving date. Cows were individually fed ad libitum total mixed rations composed of grass silage, corn silage, and concentrate during lactation and of corn silage, barley straw, and concentrate during the dry period. The metabolome was characterized in skeletal muscle samples (M. longissimus dorsi) collected on wk -7 (9 d before dry-off), -5 (6 d after dry-off), and wk -1, and 1 relative to calving. The targeted metabolomics approach was conducted using the MxP Quant 500 kit (Biocrates Life Sciences AG) with liquid chromatography, flow injection, and electrospray ionization triple quadrupole mass spectrometry. Statistical analysis on the muscle metabolite data was performed using MetaboAnalyst 5.0, which allowed us to conduct various multivariate analyses such as principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), informative heat map generation, and hierarchical clustering. The statistical analysis revealed a clear separation between pregnancy (wk -7, -5, and -1) and post-calving (wk 1). Starting 5 wk before calving and continuing through the first wk thereafter, the concentration of 3-methylhistidine (3-MH) in the muscle increased. This coincided with an increase in the concentrations of 11 AA (Phe, His, Tyr, Trp, Arg, Asn, Leu, Ile, Gly, Ser, and Thr) in the first wk after calving, whereas Gln decreased. l-arginine pathway metabolites (homoarginine, ornithine, citrulline, and asymmetric dimethylarginine), betaine, and sarcosine followed a similar pattern, increasing from wk -7 to -5, but decreasing from wk -1 to 1. The transition from pregnancy to lactation was associated with an increase in concentrations of the long-chain acylcarnitine species C16, C16:1, C18, and C18:1 in the muscle, whereas the concentrations of phosphatidylcholine and sphingomyelin in the muscle remained stable. The significant changes observed in the metabolome mainly concerned the AA and AA-related metabolites, indicating muscle protein breakdown in the first wk after calving. The metabolites produced by the L-Arg pathway might contribute to regulating skeletal muscle mass and function in periparturient dairy cows. The elevated concentrations of long-chain acylcarnitine species in the muscle in the first wk after calving suggest incomplete fatty acid oxidation, likely due to insufficient metabolic adaptation in response to the fatty acid load around the time of calving.

Novel Fatty Acid Biomarkers in Psoriasis and the Role of Modifiable Factors: Results from the METHAP Clinical Study.

Psoriasis is a chronic, immune-mediated skin condition with significant metabolic complications. Although lipid metabolism is linked to its pathogenesis, reliable biomarkers and the impact of modifiable factors remain underexplored. The aim of the present study was to identify potential biomarkers, study the affected metabolic networks, and assess the role of dietary and lifestyle factors in psoriasis. Plasma samples from 56 patients with psoriasis and 49 healthy controls were analyzed, as part of the Metabolic Biomarkers in Hashimoto's Thyroiditis and Psoriasis (METHAP) clinical trial. Using Gas Chromatography-Mass Spectrometry 23 fatty acids and their ratios were quantified, revealing significant changes in psoriasis. Specifically, lower levels of α-linoleic acid (C18:3n3), linoleic acid (C18:2n6), and gamma-linolenic acid (C18:3n6) were observed along with higher levels of eicosatrienoic acid (C20:3n3), eicosapentaenoic acid (C20:5n3), and erucic acid (C22:1n9). Total polyunsaturated fatty acids (PUFA) were significantly decreased, and the ratio of saturated to total fatty acids (SFA/Total) was increased in psoriasis (p-values < 0.0001). Linear regression identified α-linoleic acid, linoleic acid, eicosatrienoic acid, and eicosapentaenoic acid as potential biomarkers for psoriasis, adjusting for demographic, dietary, and lifestyle confounders. Network analysis revealed key contributors in the metabolic reprogramming of psoriasis. These findings highlight the association between psoriasis and fatty acid biomarkers of inflammation, insulin resistance and micronutrients deficiency, suggesting their potency in disease management.

Insight into flavor difference of cherry (Prunus avium L.) grown in facility environment and outdoors through metabolomics and correlation analysis.

The flavor profiles of cherries cultivated in greenhouse and those grown in open fields show significant variations, however, the underlying flavor-contributing factors remain unidentified. Hence, a joint investigation with widely targeted metabolomics analysis, volatile fingerprint analysis, and descriptive sensory analysis for the Russia 8 and Tieton cherry cultivars was conducted using UPLC-MS/MS and GC × GC-TOFMS to clarify the flavor differences of open-air and greenhouse-grown cherries. The study found that open-air cultivation could lead to the accumulation of non-volatile flavor substances and prompted appearance of higher acidity, astringency, plum-like flavor, and fresh herb notes; most of differential metabolites were significantly positively correlated with astringency, plum-like flavor and bitterness. Through correlation analysis and path analysis, potential flavor components and key important pathways contributing to flavor disparities were provided, and light intensity, soil moisture content, temperature and humidity were inferred as the main factors affecting the flavor profiles of open-air and greenhouse-grown cherries.

Metabolite profiling of camel milk and the fermentation bacteria agent TR1 fermented two types of sour camel milk using LC-MS in relation to their probiotic potentials.

Camel milk is a nutrient-rich diet and fermentation affects its nutritional value and probiotic function. In this study, sour camel milk and oat jujube sour camel milk were prepared using fermentation bacteria agent TR1, and the metabolites of camel milk, sour camel milk and oat jujube sour camel milk were detected using a non-targeted metabolomics approach using liquid chromatography-mass spectrometry (LC-MS).The results showed that the partial least squares discriminant analysis (PLS-DA) with 100 % accuracy and good predictive power detected 343 components in positive ion mode and 220 components in negative ion mode. The differential metabolites were mainly organic acids, amino acids, esters, vitamins and other substances contained in camel milk.It showed that there were significant differences in the metabolites of camel milk, sour camel milk and oat jujube sour camel milk. Based on the pathway enrichment analysis of the three dairy products in the KEGG database, 12 metabolic pathways mainly involved in the positive ion mode and 20 metabolic pathways mainly involved in the negative ion mode were identified. The main biochemical metabolic pathways and signal transduction pathways of the differential metabolites of the three dairy products were obtained. This study provides theoretical support for improving the nutritional quality and probiotic function of camel milk and fermented camel milk products and provides a basis for the development of relevant processing technologies and products for camel milk and fermented camel milk.

Comprehensive investigation on the flavor difference in five types of tea from JMD (Camellia sinensis 'Jinmudan').

BACKGROUND: Jinmudan (JMD) is a high-aroma variety widely cultivated in China. The current study primarily focuses on the key volatile metabolites in JMD black and oolong teas, and investigates the impact of processing technologies on the aroma quality of JMD tea. However, few studies have explored the suitability of JMD for producing a certain type of tea or the characteristic quality differences among various JMD teas using multivariate statistical analysis methods. RESULTS: The principal volatile metabolites contributing to the floral quality of JMD tea are linalool, geraniol, indole and phenethyl alcohol. In JMD black tea (BT), the key volatile metabolites include methyl salicylate, geraniol, (E)-ß-ocimene and phenethyl alcohol. In JMD oolong tea (OT), the key volatile metabolites include indole, linalyl valerate and phenethyl alcohol. In JMD yellow tea (YT), the key volatile metabolites include methyl salicylate, geraniol and terpinolene. In JMD white tea (WT), the key volatile metabolites include methyl salicylate, geraniol and terpinolene. In JMD green tea (GT), the key volatile metabolites include (E)-ß-ocimene, indole and geraniol. Comparative analysis and KEGG pathway enrichment analysis revealed that flavonoid biosynthesis is the primary metabolic pathway responsible for the taste differences among various tea types. GT exhibited higher levels of phloretin, dihydromyricetin and galangin. The contents of vitexin, tricetin in YT were relatively higher. The contents of aromadendrin and naringenin in BT were higher, while OT contained higher levels of kaempferol. Additionally, WT showed higher contents of 3-O-acetylpinobanksin and 3,5,7-pinobanksin. CONCLUSION: This study explained the reasons for the quality differences of different JMD tea and provided a reliable theoretical basis for the adaptability of JMD tea. © 2024 Society of Chemical Industry.