Untargeted Liquid Research Articles

Identification of novel biomarkers in the early diagnosis of malignant melanoma by untargeted liquid chromatography coupled to high-resolution mass spectrometry-based metabolomics: a pilot study.

Malignant melanoma (MM) is a highly aggressive form of skin cancer whose incidence continues to rise worldwide. If diagnosed at an early stage, it has an excellent prognosis, but mortality increases significantly at advanced stages after distant spread. Unfortunately, early detection of aggressive melanoma remains a challenge. To identify novel blood-circulating biomarkers that may be useful in the diagnosis of MM to guide patient counselling and appropriate disease management. In this study, 105 serum samples from 26 healthy patients and 79 with MM were analysed using an untargeted approach by liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS) to compare the metabolomic profiles of both conditions. Resulting data were subjected to both univariate and multivariate statistical analysis to select robust biomarkers. The classification model obtained from this analysis was further validated with an independent cohort of 12 patients with stage I MM. We successfully identified several lipidic metabolites differentially expressed in patients with stage I MM vs. healthy controls. Three of these metabolites were used to develop a classification model, which exhibited exceptional precision (0.92) and accuracy (0.94) when validated on an independent sample. These results demonstrate that metabolomics using LC-HRMS is a powerful tool to identify and quantify metabolites in bodily fluids that could serve as potential early diagnostic markers for MM.

HR/MS-based lipidome analysis of rat brain modulated by tolcapone

Lipids play key roles in the body, influencing cellular regulation, function, and signalling. Tolcapone, a potent catechol-O-methyltransferase (COMT) inhibitor described to enhance cognitive performance in healthy subjects, was previously shown to impact fatty acid β-oxidation and oxidative phosphorylation. However, its impact on the brain lipidome remains unexplored. Hence, this study aimed to assess how tolcapone affects the lipidome of the rat pre-frontal cortex (PFC), a region of the brain highly relevant to tolcapone therapeutic effect, while evaluating its influence on operant behaviour. Tolcapone at 20 mg/kg was chronically administered to Wistar rats during a behavioural task and an untargeted liquid chromatography high-resolution mass spectrometry (LC-HR/MS) approach was employed to profile lipid species. The untargeted analysis identified 7227 features, of which only 33% underwent statistical analysis following data pre-processing. The results revealed an improved cognitive performance and a lipidome remodelling promoted by tolcapone. The lipidomic analysis showed 32 differentially expressed lipid species in tolcapone-treated animals (FC ≥ 1.2, p-value ≤ 0.1), and among these several triacylglycerols, cardiolipins and N-acylethanolamine (NAE 16:2) were found upregulated whereas fatty acids, hexosylceramides, and several phospholipids including phosphatidylcholines and phosphatidylethanolamines were downregulated. These preliminary findings shed light on tolcapone impact on lipid pathways within the brain. Although tolcapone improved cognitive performance and literature suggests the significance of lipids in cognition, this study did not conclusively establish that lipids directly drove or contributed to this outcome. Nevertheless, it underscores the importance of lipid modulation and encourages further exploration of tolcapone-associated mechanisms in the central nervous system (CNS).

Plasma metabolomic analysis of human hepatocellular carcinoma before and after transcatheter arterial chemoembolization.

Introduction: Hepatocellular carcinoma (HCC) is the fourth most prevalent cancer in China. Transcatheter arterial chemoembolization (TACE) is a common interventional therapy for HCC. In this study, we aimed to explore specific metabolites that can accurately predict prognosis after TACE in patients with HCC. Methods: Patients with HCC and healthy volunteers (n = 20 each) were recruited to our study; plasma samples were collected from patients before and after TACE and from healthy volunteers. Plasma samples were subjected to untargeted ultra-high performance liquid chromatography-high resolution mass spectrometry metabolomics analysis, to identify metabolites significantly associated with the prognosis of patients with HCC after TACE. Results: Orthogonal filtered partial least squares discriminant analysis confirmed significant separation of the pre-TACE, post-TACE, and healthy groups, and 34 differential metabolites were identified between the pre-TACE and post-TACE groups. KEGG analysis revealed that phenylalanine, tyrosine, and tryptophan biosynthesis pathways and the phenylalanine metabolism pathway were potentially altered in HCC genesis and during TACE. Phenylalanine and tyrosine are involved in both pathways and were increased in the pre-TACE group relative to controls, with phenylalanine further increased in the post-TACE group. Receiver operating characteristic (ROC) curve analysis indicated that PC 36:4|PC 18:2_18:2 (area under the ROC curve (AUC) = 0.798) is a potential marker for assessment of prognosis in patients with HCC after TACE. Moreover, ROC curve analysis indicated that palmitoylcarnitine (AUC = 1) is a marker with potential value for HCC diagnosis. Conclusions: Limited studies had been conducted on the detection of metabolites in the plasma of HCC patients before and after TACE. PC 36:4|PC 18:2_18:2 is a potential marker for evaluation of the therapeutic effects of TACE. This finding may be beneficial for the treatment of patients with HCC after TACE.

Leukaemia Exposure Alters the Transcriptional Profile and Function of Macrophages in the Bone Marrow Niche

These authors contributed equally to this work: Martha M. Zarou & Amy Dawson Macrophages are fundamental cells of the innate immune system. Bone marrow (BM) resident macrophages are involved in antigen presentation, phagocytosis, efferocytosis (clearing of apoptotic cells) and play a critical role in regulating haematopoietic stem cell (HSC) function. In chronic myeloid leukaemia (CML), the disease-initiating leukaemic stem cells (LSCs) modulate the BM microenvironment to support leukaemia maintenance and progression. However, whether the CML BM niche alters the function of macrophages remains unknown. Using single-cell RNA sequencing (scRNA-seq) of human BM mononuclear cells, we demonstrate heterogeneity of tissue resident macrophages, isolated from CML patients at diagnosis, which cluster separately from BM macrophages deriving from heathy individuals. As most BM macrophages at diagnosis express BCR::ABL1 and are sensitive to TKI treatment, we further investigated if Philadelphia chromosome negative (Ph -) macrophages play a major role in CML development. We applied a chimeric CML mouse model (SCLtTA/BCR::ABL1) and reveal that temporal depletion of macrophages prior to leukaemia induction (using CSF1R antagonist antibody), increases leukaemia burden and decreases survival of chimeric mice following BCR::ABL1 induction (p<0.05). Comparison of CML exposed Ph - BM macrophages to control counterparts by scRNA-seq demonstrates that the CML niche drives two unique subpopulations of immature and anti-inflammatory macrophages. Functionally, we discover that Ph - BM macrophages exposed to a CML microenvironment display reduced phagocytic and efferocytic capacity, which correlates with reduction in mitochondrial respiration (p<0.05). Mechanistically, we show that diminished efferocytosis is mediated by reduced expression of the plasma membrane fatty acid translocase CD36. Additionally, treatment of primary mouse and human BM derived macrophages (BMDM) with the irreversible CD36 inhibitor sulfosuccinimidyl oleate (SSO), or genetic deletion of CD36 in THP-1 derived macrophages, results in reduced clearance of apoptotic cells in vitro. Furthermore, CD36 deficient macrophages display decreased mitochondrial respiration, similarly to CML exposed Ph - BMDM. Through untargeted liquid chromatography-mass spectrometry approach we identified the immunomodulatory protein lactotransferrin (LTF) as differentially secreted protein by leukaemic mouse stem/progenitor cells, when compared with secretome of normal counterparts (p<0.05). Treatment with LTF replicates the effect of CML exposure on BMDM, including decreased phagocytic function of mouse BMDM and THP-1 derived macrophages (p<0.05). Furthermore, mouse/human BMDM display reduced clearance of CML apoptotic cells following LTF exposure. Notably, LTF treatment reduces CD36 transcript and protein levels on BMDM and/or THP-1 derived macrophages, which correlates with reduced oxygen consumption rate and a 40% decrease in expression of the monocytic marker CD11b during macrophage differentiation, suggesting that secreted LTF is a major contributor to the immature phenotype of CML exposed macrophages. Overall, we present novel findings that dysregulated CML microenvironment modifies BM macrophage function, including their phagocytic and efferocytic capacity. This is at least partially contributed by secretion of LTF from CML stem/progenitor cells and modulation of CD36 expression on altered BM macrophages.

A comparative UPLC-MS-based metabolomics approach for distinguishing four different cultivars of Qiai (Artemisia argyi Lévl. et Van.)

The Artemisia argyi Lévl. et Van. (A. argyi) from Qichun County, Hubei Province, China, also known as Qiai, is the most important crop in the region. The quality of Qiai can be influenced by the cultivar, which is highly associated with its metabolite components. In this study, 63 batches of Qiai samples from four cultivars, Xiangai (XA), Wujian (WJ), Qijian (QJ), and Baiai (BA), were investigated using untargeted ultra-performance liquid chromatography–mass spectrometry (UPLC-MS) and chemometrics. In total, 44 metabolites were tentatively identified by combining ultra-performance liquid chromatography time-of-flight mass spectrometry (UPLC-Q-TOF/MS) data with the UNIFI platform and its embedded Traditional Medicine Library. Combining principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and orthogonal partial least squares discriminant analysis (OPLS-DA), twelve chemical markers were found to distinguish the four cultivars, showing a high degree of cultivar variability. Five of these compounds were quantified by ultra-performance liquid chromatography triple-quadrupole mass spectrometry (UPLC-TQ-MS/MS) to obtain more accurate content information. This comprehensive study constitutes a valuable reference for distinguishing the four Qiai cultivars, as well as other cultivars, and has implications for Qiai breeding.

Cochlear metabolomics, highlighting novel insights of purine metabolic alterations in age-related hearing loss

Aging is an inevitable phase in mammals that leads to health impairments, including hearing loss. Age-related hearing loss (AHL) leads to psychosocial problems and cognitive decline in the elderly. In this study, mean thresholds of auditory brainstem responses (ABR) and distortion-product otoacoustic emissions (DPOAE) increased at multiple frequencies in aged rats (14 months old) compared to young rats (2 months old). Using untargeted ultra-high performance liquid chromatography-mass spectroscopy (LC-MS), we quantified molecular metabolic markers in the cochlea of aged rats with hearing loss. A total of 137 different metabolites were identified in two groups, highlighting several prominent metabolic pathways related to purine metabolism; glycine, serine, and threonine metabolism; arginine and proline metabolism; and pyrimidine metabolism. In addition, the beneficial effects of purine supplementation were demonstrated in a mimetic model of senescent marginal cells (MCs). Overall, altered metabolic profiling is both the cause and manifestation of pathology, and our results suggest that cellular senescence and dysfunctional cochlear metabolism may contribute to the progression of AHL. These findings are seminal in elucidating the pathophysiological mechanisms underlying AHL and serve as a basis for future clinical predictions and interventions in AHL.

Identifying Serum Metabolomic Markers Associated with Skin Disease Activity in Patients with Psoriatic Arthritis.

Psoriatic arthritis (PsA) is a chronic, systemic, immune-mediated inflammatory disease causing cutaneous and musculoskeletal inflammation that affects 25% of patients with psoriasis. Current methods for evaluating PsA disease activity are not accurate enough for precision medicine. A metabolomics-based approach can elucidate psoriatic disease pathogenesis, providing potential objective biomarkers. With the hypothesis that serum metabolites are associated with skin disease activity, we aimed to identify serum metabolites associated with skin activity in PsA patients. We obtained serum samples from patients with PsA (n = 150) who were classified into mild, moderate and high disease activity groups based on the Psoriasis Area Severity Index. We used solid-phase microextraction (SPME) for sample preparation, followed by data acquisition via an untargeted liquid chromatography-mass spectrometry (LC-MS) approach. Disease activity levels were predicted using identified metabolites and machine learning algorithms. Some metabolites tentatively identified include eicosanoids with anti- or pro-inflammatory properties, like 12-Hydroxyeicosatetraenoic acid, which was previously implicated in joint disease activity in PsA. Other metabolites of interest were associated with dysregulation of fatty acid metabolism and belonged to classes such as bile acids, oxidized phospholipids, and long-chain fatty acids. We have identified potential metabolites associated with skin disease activity in PsA patients.

Genome-wide association studies identify loci controlling specialized seed metabolites in Arabidopsis.

Plants synthesize specialized metabolites to facilitate environmental and ecological interactions. During evolution, plants diversified in their potential to synthesize these metabolites. Quantitative differences in metabolite levels of natural Arabidopsis (Arabidopsis thaliana) accessions can be employed to unravel the genetic basis for metabolic traits using genome-wide association studies (GWAS). Here, we performed metabolic GWAS on seeds of a panel of 315 A. thaliana natural accessions, including the reference genotypes C24 and Col-0, for polar and semi-polar seed metabolites using untargeted ultra-performance liquid chromatography-mass spectrometry. As a complementary approach, we performed quantitative trait locus (QTL) mapping of near-isogenic introgression lines between C24 and Col-0 for specific seed specialized metabolites. Besides common QTL between seeds and leaves, GWAS revealed seed-specific QTL for specialized metabolites, indicating differences in the genetic architecture of seeds and leaves. In seeds, aliphatic methylsulfinylalkyl and methylthioalkyl glucosinolates associated with the ALKENYL HYDROXYALKYL PRODUCING loci (GS-ALK and GS-OHP) on chromosome 4 containing alkenyl hydroxyalkyl producing 2 (AOP2) and 3 (AOP3) or with the GS-ELONG locus on chromosome 5 containing methylthioalkyl malate synthase (MAM1) and MAM3. We detected two unknown sulfur-containing compounds that were also mapped to these loci. In GWAS, some of the annotated flavonoids (kaempferol 3-O-rhamnoside-7-O-rhamnoside, quercetin 3-O-rhamnoside-7-O-rhamnoside) were mapped to transparent testa 7 (AT5G07990), encoding a cytochrome P450 75B1 monooxygenase. Three additional mass signals corresponding to quercetin-containing flavonols were mapped to UGT78D2 (AT5G17050). The association of the loci and associating metabolic features were functionally verified in knockdown mutant lines. By performing GWAS and QTL mapping, we were able to leverage variation of natural populations and parental lines to study seed specialized metabolism. The GWAS data set generated here is a high-quality resource that can be investigated in further studies.

Tandem mass spectrometry (MS/MS) molecular networking guided profiling of small molecules from Aquilaria sinensis (Lour.) Gilg leaves and their bioactivity evaluation.

Agarwood, a fragrant resinous wood mainly formed by Aquilaria spp., is used worldwide as a natural fragrance and traditional medicine. A large amount of Aquilaria sinensis (Lour.) Gilg leaves are underutilised during the process of the agarwood industry, and the development of A. sinensis leaves as tea has recently attracted more and more attention. However, the small molecule profile of A. sinensis leaves and their bioactivities has been rarely reported. To conduct a rapid untargeted liquid chromatography-mass spectrometry (LC-MS) analysis of A. sinensis leaves with a molecular networking (MN) strategy and evaluate its antioxidant and antidiabetic value. A MN-assisted tandem mass spectrometry (MS/MS) analysis strategy was used to investigate the small molecule profile of A. sinensis leaves. Additionally, the integration of antioxidant and α-glucosidase inhibitory assays with MN analysis was executed to expeditiously characterise the bioactive compounds for potential prospective application. Five main chemical groups including phenol C-glycosides, organic acids, 2-(2-phenylethyl) chromones, benzophenone O-glycosides and flavonoids were rapidly revealed from the A. sinensis leaves. Eighty-one compounds were provisionally identified by comparing their MS/MS fragments with canonical pathways. The featured xanthone C-glycosides and benzophenone C-glycosides were recognised as the primary components of A. sinensis leaves. Several dimers and a trimer of mangiferin were reported firstly in A. sinensis leaves. Furthermore, 17 and 14 potential bioactive molecules were rapidly annotated from antioxidant and α-glucosidase inhibitory fraction, respectively. Our findings will help expand the utilisation of A. sinensis leaves and thus promote the high-quality development of agarwood industry.

Multimodal Imaging of Pancreatic Cancer Microenvironment in Response to an Antiglycolytic Drug.

Lipid metabolism and glycolysis play crucial roles in the progression and metastasis of cancer, and the use of 3-bromopyruvate (3-BP) as an antiglycolytic agent has shown promise in killing pancreatic cancer cells. However, developing an effective strategy to avoid chemoresistance requires the ability to probe the interaction of cancer drugs with complex tumor-associated microenvironments (TAMs). Unfortunately, no robust and multiplexed molecular imaging technology is currently available to analyze TAMs. In this study, the simultaneous profiling of three protein biomarkers using SERS nanotags and antibody-functionalized nanoparticles in a syngeneic mouse model of pancreatic cancer (PC) is demonstrated. This allows for comprehensive information about biomarkers and TAM alterations before and after treatment. These multimodal imaging techniques include surface-enhanced Raman spectroscopy (SERS), immunohistochemistry (IHC), polarized light microscopy, second harmonic generation (SHG) microscopy, fluorescence lifetime imaging microscopy (FLIM), and untargeted liquid chromatography and mass spectrometry (LC-MS) analysis. The study reveals the efficacy of 3-BP in treating pancreatic cancer and identifies drug treatment-induced lipid species remodeling and associated pathways through bioinformatics analysis.

Inulin supplementation increases the differential metabolites and metabolic pathway in Baird's tapirs (Tapirus bairdii).

The prevalence of prebiotic has increased substantially over the past decades. Little is known on its metabolic effects in zoo herbivores. We investigated the difference in faecal metabolites to characterize the composition and pathways involved after feeding inulin in zoo Baird's tapirs (Tapirus bairdii). Faecal samples were collected from before inulin treatment group and after treatment groups in six adult tapirs and analysed using untargeted liquid chromatography-mass spectrometry methods. The differential metabolites identified and metabolic pathways involved were analysed using KEGG annotation. The results demonstrated significant alterations in faecal metabolites and metabolic pathways in comparison to the control group. The amounts of differential metabolites and metabolic pathways tended to enrich with time after the treatment. We found that tryptophan and purine metabolism were relevant to the important metabolic pathways of the metabolite differences. The findings suggest that inulin may have potential applications in captive wildlife, which may provide insights into the effects of prebiotic supplementation on gut metabolites and highlights further research in this field.

Determining the role of novel metabolic pathways in driving intracranial pressure reduction after weight loss.

Idiopathic intracranial hypertension, a disease classically occurring in women with obesity, is characterized by raised intracranial pressure. Weight loss leads to the reduction in intracranial pressure. Additionally, pharmacological glucagon-like peptide-1 agonism reduces cerebrospinal fluid secretion and intracranial pressure. The potential mechanisms by which weight loss reduces intracranial pressure are unknown and were the focus of this study. Meal stimulation tests (fasted plasma sample, then samples at 15, 30, 60, 90 and 120 min following a standardized meal) were conducted pre- and post-bariatric surgery [early (2 weeks) and late (12 months)] in patients with active idiopathic intracranial hypertension. Dynamic changes in gut neuropeptides (glucagon-like peptide-1, gastric inhibitory polypeptide and ghrelin) and metabolites (untargeted ultra-high performance liquid chromatography-mass spectrometry) were evaluated. We determined the relationship between gut neuropeptides, metabolites and intracranial pressure. Eighteen idiopathic intracranial hypertension patients were included [Roux-en-Y gastric bypass (RYGB) n = 7, gastric banding n = 6 or sleeve gastrectomy n = 5]. At 2 weeks post-bariatric surgery, despite similar weight loss, RYGB had a 2-fold (50%) greater reduction in intracranial pressure compared to sleeve. Increased meal-stimulated glucagon-like peptide-1 secretion was observed after RYGB (+600%) compared to sleeve (+319%). There was no change in gastric inhibitory polypeptide and ghrelin. Dynamic changes in meal-stimulated metabolites after bariatric surgery consistently identified changes in lipid metabolites, predominantly ceramides, glycerophospholipids and lysoglycerophospholipids, which correlated with intracranial pressure. A greater number of differential lipid metabolites were observed in the RYGB cohort at 2 weeks, and these also correlated with intracranial pressure. In idiopathic intracranial hypertension, we identified novel changes in lipid metabolites and meal-stimulated glucagon-like peptide-1 levels following bariatric surgery which were associated with changes in intracranial pressure. RYGB was most effective at reducing intracranial pressure despite analogous weight loss to gastric sleeve at 2 weeks post-surgery and was associated with more pronounced changes in these metabolite pathways. We suggest that these novel perturbations in lipid metabolism and glucagon-like peptide-1 secretion are mechanistically important in driving a reduction in intracranial pressure following weight loss in patients with idiopathic intracranial hypertension. Therapeutic targeting of these pathways, for example with glucagon-like peptide-1 agonist infusion, could represent a therapeutic strategy.

Comprehensive comparison of different parts of Paeonia ostii, a food-medicine plant, based on untargeted metabolomics, quantitative analysis, and bioactivity analysis.

Paeonia ostii T. Hong & J.X. Zhang (s.s.) (Chinese name, Fengdan) is a widely cultivated food-medicine plant in China, in which root bark, seed kernels, and flowers are utilized for their medicinal and edible values. However, other parts of the plant are not used efficiently, in part due to a poor understanding of their chemical composition and potential biological activity. Untargeted ultra-performance liquid chromatography-quadrupole time of flight-mass spectrometry (UPLC-Q-TOF-MS) metabolomics was applied to characterize the metabolic profiles of 10 different parts of P. ostii. A total of 160 metabolites were alternatively identified definitely or tentatively, which were significantly different in various plant parts by multivariate statistical analysis. Quantitative analysis showed that underutilized plant parts also contain many active ingredients. Compared with the medicinal part of root bark, the root core part still contains a higher content of paeoniflorin (17.60 ± 0.06 mg/g) and PGG (15.50 ± 2.00 mg/g). Petals, as an edible part, contain high levels of quercitrin, and stamens have higher methyl gallate and PGG. Unexpectedly, the ovary has the highest content of methyl gallate and rather high levels of PGG (38.14 ± 1.27 mg/g), and it also contains surprisingly high concentrations of floralalbiflorin I. Paeoniflorin (38.68 ± 0.76 mg/g) is the most abundant in leaves, and the content is even higher than in the root bark. Branches are also rich in a variety of catechin derivatives and active ingredients such as hydrolyzable tannins. Seed kernels also contain fairly high levels of paeoniflorin and albiflorin. Fruit shells still contain a variety of components, although not at high levels. Seed coats, as by-products removed from peony seeds before oil extraction, have high contents of stilbenes, such as trans-gnetin H and suffruticosol B, showing significant potential for exploitation. Except for the seed kernels, extracts obtained from other parts exhibited good antioxidant activity in DPPH, ABTS, and ferric ion reducing antioxidant power (FRAP) assays (0.09-1.52 mmol TE/g). Five compounds (gallic acid, PGG, trans-resveratrol, kaempferol, and quercitrin) were important ingredients that contributed to their antioxidant activities. Furthermore, P. ostii seed cakes were first reported to possess agonistic activity toward CB1/CB2 receptors. This study provides a scientific basis for the further development and utilization of P. ostii plant resources.

Carnitine metabolites and cognitive improvement in patients with schizophrenia treated with olanzapine: a prospective longitudinal study.

Objective: Cognitive impairment is one of the core symptoms of schizophrenia, which is stable and lifelong. L-carnitine has been shown to improve cognitive function and decrease the rate of cognitive deterioration in patients with Alzheimer's disease. However, it remains unclear regarding the role of L-carnitine and its metabolites in cognitive functions in schizophrenia after treatment with olanzapine. The purpose of this study was to evaluate the relationship between changes in plasma levels of L-carnitine metabolites and cognitive improvement after olanzapine treatment. Methods: This was a prospective longitudinal study. In this study, we recruited 25 female patients with first episode schizophrenia (FES) who were drug naïve at baseline and received 4 weeks of olanzapine monotherapy. Cognitive function was assessed at baseline and 4-week follow-up using the RBANS. Plasma L-carnitine metabolite levels were determined by a metabolomics technology based on untargeted ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). Results: We found that the immediate memory index, delayed memory index and RBANS composite score were significantly increased at the 4-week follow-up after treatment. A total of 7 differential L-carnitine metabolites were identified in FES patients after olanzapine monotherapy. In addition, we found that changes in butyrylcarnitine were positively correlated with improvements in language index and RBANS composite score. Further regression analyses confirmed the association between reduced butyrylcarnitine levels and cognitive improvement after olanzapine monotherapy in FES patients. Conclusion: Our study shows that cognitive improvement after olanzapine treatment was associated with changes in L-carnitine metabolite levels in patients with FES, suggesting a key role of L-carnitine in cognition in schizophrenia.

LC-MS guided isolation of N,β-glucopyranosyl vincosamide and other compounds from the curare ingredient Strychnos peckii

Strychnos peckii (Loganiaceae) is an important active ingredient in curare poisons in the Amazon rainforest. Although previous studies have identified this species as a promising source of monoterpene indole alkaloids (MIAs), knowledge about other natural products is still scarce. Thus, to detect and guide the isolation of unprecedented bioactive compounds from the leaves of S. peckii, an untargeted high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) analysis was performed with the leaf aqueous extract. The HPLC-MS analysis allowed the detection of eleven compounds, including the alkaloids harman-3-carboxylic acid (5) and N,β-glucopyranosyl vincosamide (6), and the flavonoids quercetin 3-O-rhamnopyranoside (9) and kaempferol 3-O-rhamnopyranoside (10), all not previously reported in the Loganiaceae family. These compounds, along with strictosidine (3), 5-carboxystrictosidine (7), and desoxycordifoline (8) were isolated through modern chromatographic techniques and determined by using NMR spectroscopy in combination with MS. Overall, the untargeted HPLC-MS analysis proved to be a simple and effective approach to guide the isolation of substances not yet identified from S. peckii.

Metabolomics Profiling of White Button, Crimini, Portabella, Lion's Mane, Maitake, Oyster, and Shiitake Mushrooms Using Untargeted Metabolomics and Targeted Amino Acid Analysis.

Mushrooms contain multiple essential nutrients and health-promoting bioactive compounds, including the amino acid L-ergothioneine. Knowledge of the chemical composition of different mushroom varieties will aid research on their health-promoting properties. We compared the metabolomes of fresh raw white button, crimini, portabella, lion's mane, maitake, oyster, and shiitake mushrooms using untargeted liquid chromatography mass spectrometry (LC/MS)-based metabolomics. We also quantified amino acid concentrations, including L-ergothioneine, a potential antioxidant which is not synthesized by plants or animals. Among the seven mushroom varieties, more than 10,000 compounds were detected. Principal Component Analysis indicated mushrooms of the same species, Agaricus Bisporus (white button, portabella, crimini), group similarly. The other varieties formed individual, distinct clusters. A total of 1344 (520 annotated) compounds were detected in all seven mushroom varieties. Each variety had tens-to-hundreds of unique-to-mushroom-variety compounds. These ranged from 29 for crimini to 854 for lion's mane. All three Agaricus bisporus varieties had similar amino acid profiles (including detection of all nine essential amino acids), while other varieties had less methionine and tryptophan. Lion's mane and oyster mushrooms had the highest concentrations of L-ergothioneine. The detection of hundreds of unique-to-mushroom-variety compounds emphasizes the differences in chemical composition of these varieties of edible fungi.

Identification of altered metabolic functional components using metabolomics to analyze the different ages of fruiting bodies of Sanghuangporus vaninii cultivated on cut log substrates.

Sanghuangporus vaninii is a profitable traditional and medicinal edible fungus with uncommon therapeutic properties and medicinal value. The accumulation of active ingredients in this fungus that is used in traditional Chinese medicine is affected by its years of growth, and their pharmacological activities are also affected. However, the effects of age on the medicinal value of fruiting bodies of S. vaninii cultivated on cut log substrate remain unclear. In this study, an untargeted liquid chromatography mass spectrometry (LC-MS)-based metabolomics approach was performed to characterize the profiles of metabolites from 1-, 2- and 3-year-old fruiting bodies of S. vaninii. A total of, 156 differentially accumulated metabolites (DAMs) were screened based on the criterion of a variable importance projection greater than 1.0 and p < 0.01, including 75% up regulated and 25% down regulated. The results of enrichment of metabolic pathways showed that the metabolites involved the biosynthesis of plant secondary metabolites, biosynthesis of amino acids, central carbon metabolism in cancer, steroid hormone biosynthesis, linoleic acid metabolism, prolactin signaling pathway, and arginine biosynthesis, and so on. The biosynthesis of plant secondary metabolites pathway was significantly activated. Five metabolites were significantly elevated within the growth of fruiting bodies, including 15-keto-prostaglandin F2a, (4S, 5R)-4,5,6-trihydroxy-2-iminohexanoate, adenylsuccinic acid, piplartine, and chenodeoxycholic acid. 15-keto-prostaglandin F2a is related to the pathway of arachidonic acid metabolism and was significantly increased up to 1,320- and 535-fold in the 2- and 3-year-old fruiting bodies, respectively, compared with those in the 1-year-old group. The presence of these bioactive natural products in S. vaninii is consistent with the traditional use of Sanghuang, which prompted an exploration of its use as a source of natural prostaglandin in the form of foods and nutraceuticals. These findings may provide insight into the functional components of S. vaninii to develop therapeutic strategies.

The inhibition effects of chlorogenic acid on the formation of colored oxidation products of (-)-epigallocatechin gallate under enzymatic oxidation.

Untargeted Liquid chromatography tandem mass spectrometry (LC-MS) based metabolomics in combination with UV-visible and colorimeter was applied in identifying critical colored enzymatically oxidized products of (-)-epigallocatechin gallate (EGCG). Pearson correlation coefficient analysis between marker compounds and a* value was conducted, and then a series of colored oxidation products were targeted and subsequently identified by diode array detection and mass fragmentation ions. The quinone of oolongtheanin 3-O'-gallate degraded product with quasi-molecular mass ion at m/z 711 was identified as a critical colored oxidation product of single EGCG. To explore the effect of chlorogenic acid on the formation of colored EGCG enzymatic oxidation products, the variation of oxidation products on the oolongtheanin pathway was semi-quantitatively determined. The result showed chlorogenic acid significantly inhibited the formation of colored oxidation products, thus lightened the color of EGCG oxidation mixture. The addition of chlorogenic acid influences the process of tea polyphenols' enzymatic oxidation.

Untargeted metabolomics liquid chromatography-high resolution mass spectrometry approach for the geographical origin assessment of Italian dehydrated apples.

Geographical provenience is nowadays a relevant aspect of the authenticity and the quality of many food commodities. Dehydrated apple cubes/slices represent an ingredient commonly used by food companies for bakery products. However, this apple-based matrix is not so known and studied from an analytical point of view. In the present work, seven compounds were identified as key molecules to distinguish between Italian and non-Italian samples, through an untargeted ultrahigh-pressure liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) approach. This methodology was merged with multivariate statistical analysis, and the principal features were studied and identified considering several identification steps. Samples from 2020 and 2021 harvesting campaigns, with partial and total dehydration rates, with or without peel, and from different apple varieties were considered for the study, for a total of 91 samples. Afterward, the same analysis protocol was applied to an external set (n = 12 samples), included in the statistical models, searching for the key compounds identified in the training set. Interesting and significant results underlined the potentiality of the UHPLC-HRMS technology as a confirmatory strategy for the geographical origin assessment of dehydrated apple commodities.

Unexplored Potential: Metabolite Screening of Local Lake Algae Isolated from Al-Asfar Lake in Saudi Arabia

In this study, we aimed to explore the commercial potential of various microalgae variants found in Al-Asfar Lake, Saudi Arabia, which have not been extensively investigated previously. We conducted a comprehensive analysis of the metabolic profiles of algae isolated from Al-Asfar Lake. The isolated algae were subjected to molecular analysis using specific primers for the Chlorophyceae class to confirm their identity. Subsequently, we compared the concentration of metabolites in the locally isolated Chlorella vulgaris from Al-Asfar Lake with five commercially available algae (Tetraselmis, Nannochloropsis, Chlorella vulgaris, Spirulina, and Isochrysis). To perform the metabolomics analysis, we employed untargeted ultra-performance liquid chromatography (UPLC) coupled with mass spectrometry (MS) analysis, which yielded a total of 168 metabolites from the microalgae samples. The data were further analyzed using MetaboAnalyst. The results revealed two distinct clusters of microalgae: the first cluster comprised Chlorella vulgaris and the microalgae isolated from the lake, while the second cluster consisted of two sub-clusters, with Isochrysis grouped with Tetraselmis, and Nannochloropsis clustered with Spirulina. Notably, the metabolites of Al-Asfar Lake algae showed a remarkable similarity to Chlorella vulgaris. These findings have significant implications for the environmental aspect of Al-Asfar Lake, shedding light on critical insights into the metabolites and commercial potential of the lake’s microalgae. The valuable insights gained from this research can be utilized to investigate the impact of nutrient abundance on the lake’s biodiversity, enhance microalgal biomass production for biofuel applications, and explore the reuse of lake water in agriculture and environmental restoration projects. Overall, our study provides important groundwork for understanding the potential of Al-Asfar Lake microalgae and their application in various industries, contributing to the sustainable development and environmental health of the region.