Ultra-high Performance Liquid Chromatography-mass Spectrometry Research Articles

Metabolomic analysis of HUVEC after Thermal denaturation UHPLC-MS/MS-based metabolomics.

Preserving denatured dermis has been shown to promote wound healing and improve skin appearance and function. Angiogenesis is crucial for the healing of burn wounds. However, the metabolic mechanisms underlying angiogenesis during burn recovery remain unclear. In this study, ultra-high performance liquid chromatography-mass spectrometry analysis revealed six distinct metabolites in a heat-denatured cell model. A bioinformatics approach was used to predict the differentially expressed metabolites, and four metabolic pathways closely related to trauma repair were identified. These pathways might play a significant role in the regression of thermally injured endothelial cells. We also found that increasing D-mannose level promoted the angiogenic activity of human umbilical vein endothelial cells in the heat-denatured cell model, enhancing cell proliferation, migration, and tube formation. In summary, these findings revealed changes in metabolites and metabolic pathways in thermally injured endothelial cells, and demonstrated that D-mannose could promote angiogenesis during the recovery of thermally injured endothelial cells.

Utilization and validation of dried blood spot-based metabolomics in plasma-derived diagnostic models

Metabolomics has rapidly advanced in life sciences, enhancing our understanding of physiological conditions. Plasma stands out as a predominant sample type in metabolomics studies. Dried blood spot (DBS) has been recognized as a valuable strategy due to its unique characteristics, such as minimal invasiveness, small sample volume, and easy transport. Most large cohort studies construct the diagnostic model using plasma-based metabolomics rather than DBS. However, the comparability of metabolite measurements between DBS and plasma samples, as well as the integration of DBS-based metabolomics into established plasma-derived diagnostic models, remains unclear. We collected plasma and DBS samples from the same 12 healthy controls under fasting and non-fasting conditions and performed ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS)-based metabolomics. We identified 277 metabolites present in both plasma and DBS samples, with 229 (82.7 %) showing a strong correlation between the two sample types. Furthermore, pre-processing datasets from plasma and DBS samples before combining them effectively diminished inconsistencies between plasma- and DBS-based metabolomics without compromising the inherent classification within the data. For the clinical application, the plasma samples were obtained from the gastric cancer patients (n = 204) and the healthy controls (n = 128) to serve as the training cohorts. Additionally, DBS samples were collected from different participants of gastric cancer (n = 19) and healthy controls (n = 12) to validate the diagnostic model. Finally, DBS-based metabolomics were integrated into established plasma-derived gastric cancer diagnostic models, yielding an area under the ROC curve (AUC) of 0.934. Overall, incorporating DBS-based metabolomics into an established plasma-derived diagnostic model holds promise, offering valuable opportunities and insights for diagnostic research and clinical practice.

The use of metabolomics and machine learning algorithms to predict post-transplant diabetes mellitus in renal transplant patients on Tacrolimus therapy.

Tacrolimus (TAC) has significantly improved kidney graft survival following transplantation, though it is associated with adverse side effects. The most prevalent complication resulting from excessive TAC exposure is the onset of de novo diabetes mellitus (DM), a condition that can negatively impact both renal graft function and patient outcomes. De novo DM is linked to an increased risk of chronic transplant dysfunction, as well as cardiovascular morbidity and mortality. Although the underlying mechanisms remain unclear, emerging research in the field of omics shows promise. The aim of this study was to investigate the metabolomic profile of kidney transplant patients who developed de novo DM, in comparison to those who did not, following TAC exposure, using untargeted metabolomic analysis through ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS) and machine learning algorithms. A cohort of 34 kidney transplant patients on a Tacrolimus regimen for at least 6 months was enrolled in the study, with serum samples collected from each patient. Comprehensive profiling of serum metabolites was performed, enabling the classification of patients into de novo diabetes mellitus and non diabetes groups. The metabolomic analysis of serum was conducted using UHPLC-MS. Of the 34 patients, 16 were diagnosed with TAC-induced diabetes. A total of 334 metabolites were identified in the serum samples, of which 10 demonstrated a significant correlation with the de novo diabetes mellitus group. Most of these metabolites were linked to alterations in lipid metabolism. The application of metabolomics in kidney transplant patients undergoing a Tacrolimus regimen is both feasible and effective in identifying metabolites associated with de novo diabetes mellitus. This approach may provide valuable insights into the metabolic alterations underlying TAC-induced diabetes.

Analysis of Azathioprine Metabolites in Autoimmune Hepatitis Patient Blood-Method Development and Validation.

Autoimmune hepatitis (AIH) is a chronic inflammatory liver disease treated by steroids and immunomodulator thiopurine drugs such as azathioprine (AZA). AZA is metabolized in the human body into bioactive forms such as 6-thioguanine (6-TG) and 6-methyl-mercaptopurine (6-MMP). Monitoring the levels of bioactive AZA metabolites is very important for proper treatment of patients. In this study, our aim was to develop and validate a fast and sensitive ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) method for the analysis of 6-TG and 6-MMP from blood samples of patients with AIH to monitor the level of these bioactive metabolites. The detection and quantification of the analytes was carried out by Selected Reaction Monitoring (SRM)-based targeted mass spectrometry. The method was validated according to the EMA guidelines. Blood samples from patients with AIH treated with AZA were analysed with the developed method. The method was successfully validated with appropriate accuracy and precision for the target biomolecules and their concentration in the samples from patients with AIH was determined. The developed and validated UHPLC-MS method enables the fast and precise analysis of AZA metabolites.

Exploring the Potential Mechanisms of Danshen for the Treatment of Ulcerative Colitis based on Serum Pharmacochemistry, Gene Expression Profiling, and Network Pharmacology: Regulation of Cell Apoptosis and Inflammatory Response.

As a traditional Chinese medicine, Danshen shows potential efficacy for treating ulcerative colitis (UC). However, the bioactive components and mode of action were unclear. This paper uses a combination of network pharmacology, serum medicinal chemistry, and gene expression profiling to clarify its possible molecular mechanism of action and material basis. Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) was utilized to analyze the herbal components and metabolites from the serum of Danshen-treated mice. Gene expression profiles were applied to construct a database of Danshen action targets. Then, active ingredient-target-biological functional module networks were constructed to analyze the mechanism of action. Molecular docking has further confirmed the possibility of its components to the targets. As a result, 193 common targets between 1684 Danshen-related DEGs and 1492 UC targets were determined as the potential targets for Danshen in treatment with UC. Serum pharmacochemistry and target prediction showed that 22 components in serum acted on 777 targets. Intersection with common targets yielded 46 core targets, and an active ingredienttarget- biological functional module network was constructed for analysis. Network prediction and molecular docking results showed that the main action modules were inflammatory response and cell apoptosis, which mainly acted on targets SRC, RELA, HSP90AA1, CTNNB1, STAT3, and CASP3. The main components of Danshen intervention in UC were predicted to include Catechol, 3,9-Dimethoxypterocarpan, 8-Prenylnaringenin, Isoferulic acid, Salvianolic acid C, and Danshensu. The present study provides a scientific foundation for further explicating the mechanisms of Danshen against UC.

Reversed-Phase Ultrahigh-Performance Liquid Chromatography-Mass Spectrometry Method for High-Throughput Lipidomic Quantitation.

Reversed-phase ultrahigh-performance liquid chromatography-mass spectrometry (RP-UHPLC/MS) method is optimized for the quantitation of a large number of lipid species in biological samples, primarily in human plasma and serum. The method uses a C18 bridged ethylene hybrid (BEH) column (150×2.1mm; 1.7μm) for the separation of lipids from 23 subclasses with a total run time of 25min. Lipid species separation allows the resolution of isobaric and isomeric lipid forms. A triple quadrupole mass spectrometer is used for targeted lipidomic analysis using multiple reaction monitoring (MRM) in the positive ion mode. Data are evaluated by Skyline software, and the concentrations of analytes are determined using internal standards per each individual lipid class.

Low polarity fraction of Radix Bupleuri alleviates chronic unpredictable mild stress-induced depression in rats through FXR modulating bile acid homeostasis in liver, gut, and brain

Radix Bupleuri (BR, Bupleurum chinense DC.) is a well-known traditional Chinese medicine (TCM) known for its effects on soothing the liver and alleviating depression, and is widely used in clinical settings to manage depressive symptoms. A dosage of 12.5g crude drug/kg/d of the low-polarity fraction of Radix Bupleuri (LBR) demonstrated effectiveness in treating depression in our previous study. However, the mechanism through which BR ameliorates depression remains unclear. This study aimed to explore the polar fractions of BR and their mechanisms of action in the treatment of depression. Chronic unpredictable mild stress (CUMS) rats were continuously administered BR by oral gavage for 4 weeks. Behavioral and biochemical indicators were evaluated to assess the antidepressant effects of LBR, and transcriptomics was used to explore the relevant pathways. In addition, pseudo-targeted bile acid (BA) metabonomics was used to quantify the BA profiles. Molecular biology techniques have been used to investigate the underlying mechanisms. LBR serves as a more effective active fraction with antidepressant activity. Intervention with LBR, which is characterized by a clearly defined chemical composition, significantly ameliorated depression-like behavior and biochemical indicators in rats subjected to CUMS. Notably, marked improvements were observed in the levels of total bile acids (TBAs) in the blood, liver, and ileum. Mechanistically, liver transcriptome analysis suggested that bile secretion may be a crucial pathway for alleviating depression after LBR treatment. Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) BA metabonomics indicated that TCA, β-MCA, γ-MCA, Tβ-MCA, and UDCA in the liver, Tβ-MCA, TCA, βMCA, GHDCA, and GLCA in the ileum, and β-MCA, CA, and DCA in the hippocampus were the potential therapeutic targets. In addition, molecular biology experiments showed that LBR exerts antidepressant effects by regulating the FXR/SHP/CYP7A1 pathway in the liver, the FXR/FGF15/ASBT pathway in the ileum, and the FXR/CREB/BDNF pathway in the hippocampus. In conclusion, LBR attenuated depression by moderating BA homeostasis through FXR and related genes within the liver-gut-brain axis.

Effects of Shaofu Zhuyu decoction on intestinal flora and fibrosis in a mouse model of endometriosis

Shaofu Zhuyu decoction has been widely used to treat gynecological diseases; however, its mechanism of action in endometriosis remains unclear. We analyzed Shaofu Zhuyu decoction’s chemical composition using ultra-high performance liquid chromatography-mass spectrometry. In an endometriosis mouse model, ectopic lesions weight measurements and hematoxylin and eosin staining were used to assess the therapeutic efficacy of Shaofu Zhuyu decoction. Effects on intestinal microflora were analyzed using 16S ribosomal ribonucleic acid sequencing, and impacts on focal fibrosis were analyzed using Masson’s trichrome staining. Moreover, fibrosis- and metabolism-related proteins were assessed using immunohistochemistry and enzyme-linked immunosorbent assay. The study identified 157 chemical constituents within Shaofu Zhuyu decoction. Shaofu Zhuyu decoction treatment in mice with endometriosis resulted in a reduction in ectopic lesions weight (P<0.05) and delayed disease progression. Moreover, it improved the diversity and abundance of intestinal flora, and decreased the expression of Lachnospiraceae (P<0.05), Rikenellaceae (P<0.01), Ruminococcaceae (P<0.01), Lachnoclostridium (P<0.05), and unclassified_f__Ruminococcaceae (P<0.05). Kyoto Encyclopedia of Genes and Genomes analysis revealed enrichment in carbohydrate, amino acid, and lipid metabolism pathways. Masson’s trichrome staining revealed that compared to the untreated group, the Shaofu Zhuyu decoction group exhibited significantly reduced collagen deposition areas (P<0.001), lower TGF-β1 (P<0.001), COL1A1 (P<0.05), and α-SMA (P<0.01) expression in ectopic lesions, along with increased serum adiponectin (P<0.05), decreased serum leptin (P<0.05), TGF-β1 (P<0.001), and CTGF (P<0.05). Shaofu Zhuyu decoction regulates the intestinal flora of mice with endometriosis while also reducing fibrosis at the lesion site. These findings highlight novel mechanisms for its efficacy in alleviating endometriosis.

Dissipation and Dietary Risk Assessment of the Fungicide Pyraclostrobin in Apples Using Ultra-High Performance Liquid Chromatography-Mass Spectrometry.

The fungicide pyraclostrobin is the main measure used to control apple alternaria blotch in production. To evaluate the potential dietary risks for consumers, the dissipation and terminal residues of pyraclostrobin were investigated using ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS). Pyraclostrobin in apples was extracted by acetonitrile with 2% ammonia and then purified using primary secondary amine (PSA) and graphitized carbon black (GCB). The method showed good linearity within the concentration range of 0.005-0.1 mg L-1, with a coefficient of determination (R2) ≥ 0.9958. The recoveries ranged from 96.0% to 103.8%, with relative standard deviations (RSDs) between 0.8% and 2.3%. The limit of quantification (LOQ) was 0.01 mg kg-1. Pyraclostrobin dispersible oil suspension was applied in 12 apple fields across China according to good agricultural practices (GAPs). In Beijing and Shandong, the dissipation of pyraclostrobin followed first-order kinetic equations, with a half-life of 11 days. The terminal residues ranged from <0.01 to 0.09 mg kg-1. The national estimated daily intake (NEDI) of pyraclostrobin was compared with the acceptable daily intake (ADI), resulting in risk quotient (RQc) of 80.8%. These results suggest that pyraclostrobin poses a low health risk to consumers under GAP conditions and according to recommended dosages.

Novel eicosanoid signature in plasma provides diagnostic for metabolic dysfunction-associated steatotic liver disease

There is a clinical need for a simple test implementable at the primary point of care to identify individuals with metabolic dysfunction-associated steatotic liver disease (MASLD) in the population. Blood plasma samples from adult patients with varying phenotypes of MASLD were used to identify a minimal set of lipid analytes reflective of underlying histologically confirmed MASLD. Samples were obtained from the NIDDK Nonalcoholic Steatohepatitis Clinical Research Network (NASH CRN) NAFLD Database prospective cohort study (MASLD group; N = 301). Samples of control subjects were obtained from cohort studies at the University of California San Diego (control group; N = 48). Plasma samples were utilized for targeted quantitation of circulating eicosanoids, related bioactive metabolites, and polyunsaturated fatty acids by ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) lipidomics analysis. Bioinformatic approaches were used to discover a panel of bioactive lipids that can be used as a diagnostic tool to identify MASLD. The final panel of fifteen lipid metabolites consists of 12 eicosanoid metabolites and 3 free fatty acids that were identified to be predictive for MASLD by multivariate area under the receiver operating characteristics curve (AUROC) analysis. The panel was highly predictive for MASLD with an AUROC of 0.999 (95% CI = 0.986–1.0) with only one control misclassified. A validation study confirmed the resulting MASLD LIPIDOMICS SCORE, which may require a larger-scale prospective study to optimize. This predictive model should guide the development of a non-invasive “point-of-care” test to identify MASLD patients requiring further evaluation for the presence of metabolic dysfunction-associated steatohepatitis.

Herpes simplex reactivation triggers symptom exacerbation in patients with major depressive disorder

BackgroundHerpes Simplex Virus (HSV) infection has been linked to depression, but the underlying mechanism remains largely unknown. We hypothesized that recurring HSV infection could exacerbate depressive symptomatology via activation of the immune response and the kynurenine pathway in patients with Major Depressive Disorder (MDD). Methods32 MDD patients and 17 psychiatrically healthy individuals with a history of HSV recurrence were enrolled into the study. Blood samples were obtained, and psychiatric symptom severity was assessed at three separate visits- asymptomatic, active infection, and follow-up. Plasma levels of tryptophan metabolite levels were measured using ultra-high performance liquid chromatography-mass spectrometry (LC-MS), and inflammation-associated proteins were measured using high sensitivity electrochemiluminescence. ResultsThere was a significant effect of infection recurrence on IFN-γ and sICAM-1 levels, the latter being influenced by MDD diagnosis. This indicates that the effect of infection on sICAM-1 levels differs between control and depressed groups, although post hoc analysis suggests that interaction effect might be subtle. Infection recurrence had a significant effect on the reactivity of psychiatric symptoms. Finally, there was a positive correlation between levels of anti-inflammatory cytokines IL-10 and IL-13 and depression severity in the cohort during infection recurrence. LimitationsRelatively small sample size, limited number of male participants, and the relatively weak inflammatory stimuli studied. ConclusionsMDD patients might react with an exacerbation of symptoms along with elevations of inflammatory markers during HSV recurrence. Further, the association with symptom severity indicates that therapeutic modulation of inflammation may be beneficial in patients with MDD, especially during active infections.

Anti-biofilm and anti-virulence potential of cell free supernatant of Akkermansia muciniphila against Salmonella

Akkermansia muciniphila is one of the commensals residing within the mammalian gut and co-evolving with the host. Numerous studies have demonstrated the benefits of A. muciniphila in ameliorating metabolic disorders, while little is known about the antimicrobial potential of A. muciniphila against pathogens. Here, we examined the antimicrobial and anti-virulence properties of cell free supernatant (CFS) of A. muciniphila against Salmonella Typhimurium. CFS retarded bacterial growth and inhibited the motility of S. Typhimurium SL1344 and S. Typhimurium 14028. CFS dose-dependently reduced cell hydrophobicity and auto-aggregation of both strains. Also, CFS from A. muciniphila significantly attenuated biofilm formation. Compared with untreated bacteria, CFS-treated bacteria significantly decreased adhesion and invasion to Caco-2 cells, and reduced intracellular survival in macrophages. CFS maintained antimicrobial properties after treatment with high temperatures and various proteases, while it lost its antimicrobial activity after pH neutralization. Gas chromatography-mass spectrometry (GC-MS) confirmed that A. muciniphila produced a certain amount of acetate and propionate, and ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS) identified other organic acids and metabolites in CFS. In summary, CFS from A. muciniphila exhibited anti-biofilm and anti-virulence properties against Salmonella and could be potentially utilized in the food industry for controlling Salmonella contamination and reducing infection.

Screening bioactive compounds from Fangji Huangqi decoction for treating rheumatoid arthritis via COX-2 magnetic ligand fishing combined with in vivo validation

Ethnopharmacological relevanceFangji Huangqi Decoction (FHD) is a classical Chinese compound formula for treating rheumatoid arthritis (RA) with satisfactory effects. FHD is reputed for its ability to tonify qi with strengthening exterior, and dispel wind while removing dampness, but its mechanisms and bioactive compounds for treating RA remain unclear. Aim of the studyThe aim of this study was to explore the key target and bioactive compounds that were responsible for FHD-mediated improvements in RA. Materials and methodsUsing network pharmacology, we discovered that cyclooxygenase-2 (COX-2) was the key target of FHD against RA. We utilized a ligand fishing technique with COX-2 immobilized magnetic beads to recognize the bioactive components that act on COX-2. Then we carried out an in vitro assay of COX-2 enzyme inhibition and in vivo assay of carrageenan-induced inflammation and collagen-induced arthritis (CIA) to validate the bioactive effects of these captured ingredients. In the CIA assay, micro-CT, hematoxylin‒eosin staining and safranin-O/fast green staining were employed to assess the influence of the captured ligand on bone damage, pathological injury and cartilage destruction, respectively. Immunohistochemistry (IHC) and enzyme-linked immunosorbent assays (ELISAs) were used to detect the expression of COX-2 target in the ankle joint. interleukin-6 (IL-6) levels in the serum were also detected by ELISA. Molecular docking was used to reveal the binding mechanism of the COX-2 protein and the captured ligand. ResultsEleven ligands, including tetrandrine, fangchinoline, cyclanoline, licochalcone B, ononin, calycosin and liquiritin, were specifically bound to the COX-2 protein, as determined by ultrahigh-performance liquid chromatography–mass spectrometry (UPLC–MS), seven of which were present at high levels. One ligand, tetrandrine, not only had a great inhibitory effect on COX-2 enzyme activity but also significantly reduced carrageenan-induced inflammation. In the CIA assay, middle- and high-dose tetrandrine (25 and 50 mg/kg) had effects comparable to those of FHD and celecoxib on ameliorating RA symptoms in CIA mice via the COX-2 target. Furthermore, compared with the low-dose tetrandrine group (12.5 mg/kg), the FHD group exhibited significantly lower arthritis index scores and serum IL-6 expression, although the content of tetrandrine in FHD extract solution was approximately 0.1% of that in the low-dose tetrandrine group. ConclusionsHence, we inferred that tetrandrine was the main bioactive component responsible for the effects of FHD against RA by suppressing the expression of the COX-2 protein and inhibiting the enzyme catalytic activity of COX-2. The reason for these effects may be that tetrandrine can interact with the residue Tyr385 of COX-2, the enzymatic catalytic site of COX-2 to transform arachidonic acid (AA) to prostaglandin E2 (PGE2), and thereby reduce the production of prostaglandins and inflammatory metabolites. Moreover, in addition to tetrandrine, FHD contains other compounds that could supplement the activity of tetrandrine when FHD was used to treat RA, which is manifested the “multi-component” characteristic of how Traditional Chinese Medicine formulas treat diseases.

Determination of fluoroquinolones residues in bean sprouts using covalent organic framework based solid-phase extraction coupled with UHPLC-MS/MS

A solid-phase extraction (SPE) method using three-dimensional covalent organic framework (COFs) coupled with ultra-high performance liquid chromatography-mass spectrometry was developed for the detection of 13 fluoroquinolones (FQs) in bean sprout samples. Critical extraction parameters influencing SPE efficiency were optimized and matrix-matched standard curves were used to eliminate matrix effects and validate the proposed method. The analytical method proved to have good linearity in the concentration range of 1.0–100 μg L–1 with correlation coefficients of r≥0.9995. The limits of detection were 0.10–0.45 μg kg–1, whereas the limits of quantification were 0.34–1.51 μg kg–1. The recoveries ranged from 92.7 % to 105 % with satisfactory precision (< 4.82 %). The COFs-packed SPE cartridge maintained recovery efficiency for the target FQs over five reuse cycles. The adsorption selectivity was mainly attributed to π–π interactions, hydrogen bonding, and electrostatic attraction between FQs and the adsorbent. Analysis of actual samples revealed the presence of enrofloxacin, ciprofloxacin, and norfloxacin in three batches of bean sprout samples. These results highlight the importance of monitoring antibiotics residues in bean sprouts.

Widely Targeted Metabolomics Analysis to Reveal Metabolite of Morus alba L. in Different Medicinal Parts.

Morus alba L. is a tradition medical and edible plant. It is rich in many important bioactive components. However, there is a dearth of systematic information about the components. Here, the Mori Cortex, Mori Folium, Mori Fructus, and Mori Ramulus were studied. Ultrahigh-performance liquid chromatography-mass spectrometry (UHPLC-MS) is used to study primary and secondary metabolites. Eight hundred two metabolites were identified and classified into 10 different categories in total. Correlation analysis, hierarchical clustering analysis, and principal component analysis of metabolites showed that different parts of the sample could be significantly different. In different medicinal parts, alkaloids accounted for 4.0%, 3.6%, 5.1%, and 4.5%; flavonoids accounted for 0.7%, 27.2%, 5.6%, 1.2%; terpenes accounted for 20.1%, 2.1%, 2.6%, 2.5%. Furthermore, the abundance of phenols, phenylpropanoids, and lipids metabolites sequentially accounted for 2.3-4.4%, 0.5-1.8%, and 2.4-5.3%. These results have improved our understanding of metabolites and provided a reference for research on the medicinal and edible value of Morus alba L. In addition, the study reveals the correlation between the components of Traditional Chinese medicine and the basic theory of TCM properties and reinterprets the ancient wisdom in the world's traditional herbs through the perspective of modern science.

Dysregulated arginine metabolism in precursor B-cell acute lymphoblastic leukemia in children: a metabolomic study

BackgroundPrecursor B-cell acute lymphoblastic leukemia (B-ALL) is the most common cancers in children. Failure of induction chemotherapy is a major factor leading to relapse and death in children with B-ALL. Given the importance of altered metabolites in the carcinogenesis of pediatric B-ALL, studying the metabolic profile of children with B-ALL during induction chemotherapy and in different minimal residual disease (MRD) status may contribute to the management of pediatric B-ALL.MethodsWe collected paired peripheral blood plasma samples from children with B-ALL at pre- and post-induction chemotherapy and analyzed the metabolomic profiling of these samples by ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS). Healthy children were included as controls. We selected metabolites that were depleted in pediatric B-ALL and analyzed the concentrations in pediatric B-ALL samples. In vitro, we study the effects of the selected metabolites on the viability of ALL cell lines and the sensitivity to conventional chemotherapeutic agents in ALL cell lines.ResultsForty-four metabolites were identified with different levels between groups. KEGG pathway enrichment analyses revealed that dysregulated linoleic acid (LA) metabolism and arginine (Arg) biosynthesis were closely associated with pediatric B-ALL. We confirmed that LA and Arg were decreased in pediatric B-ALL samples. The treatment of LA and Arg inhibited the viability of NALM-6 and RS4;11 cells in a dose-dependent manner, respectively. Moreover, Arg increased the sensitivity of B-ALL cells to L-asparaginase and daunorubicin.ConclusionArginine increases the sensitivity of B-ALL cells to the conventional chemotherapeutic drugs L-asparaginase and daunorubicin. This may represent a promising therapeutic approach.

Exploring the mechanism of Pueraria lobata (Willd.) Ohwi in the regulation of obesity

Ethnopharmacological relevancePueraria lobata (Willd.) Ohwi is a traditional medicinal and edible homologous plant rich in flavonoids, triterpenes, saponins, polysaccharides and other chemical components. At present, studies have shown that Pueraria lobata radix (PR) has the effect of lowering blood sugar, improving insulin sensitivity and inhibiting obesity. However, the specific mechanism of PR inhibits obesity is still unclear, and there are few researches on the anti-obesity effect of PR through the combination of network pharmacology and experiment. Aim of the studyPharmacology, molecular docking technology and experimental verification through the network, revealing the PR the material basis of obesity and the potential mechanism. Methods and resultsThe present study used network pharmacology techniques to investigate the therapeutic effect and mechanism of action of PR. Through relevant databases, a total of 6 main chemical components and 257 potential targets were screened. Protein interaction analysis shows that AKT1, AKR1B1, PPARG, MMP9, TNF, TP53, BAD, and BCL2 are core targets. Enrichment analysis shows that the pathway of PR in preventing obesity involves the cancer signaling pathway and the PI3K-Akt signaling pathway, which may be the main pathways of action. Further molecular docking verification indicates that its core target exhibits good binding activity with 4 compounds: formononectin, purerin, 7,8,4 ′- trihydroxide and daidzein. Using the ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS) technology to detected and confirmed these main compounds. Cell experiment results revealed that puerarin inhibits cell proliferation and differentiation in a concentration dependent manner, significantly promoting cell apoptosis and affecting cell migration. Animal experiments have shown that puerarin reduces food intake and weight gain in mice. It was found that puerarin can upregulate HDL and downregulate TC, TG, and LDL blood biochemical indicators. Western blot results showed that puerarin significantly inhibited the expression of AKT1, AKR1B1, MMP9, TNF, TP53, BCL2, PPARG, and significantly increased the expression of BAD protein at both cellular and animal levels. ConclusionThe present study established a method for measuring PR content and predicted its active ingredients and their mechanisms of action in the treatment of obesity, providing a theoretical basis for further research.

Antibiotic Pollution and Its Effects on the Spatiotemporal Variation in Microbial Community Structure and Functional Genes in Sediment of Baiyangdian Lake

Microbial communities play an important role in water quality regulation and biogeochemical cycles in lakes, and their community structure and function are affected by environmental factors. Therein, antibiotics affect the abundance, diversity, composition, and function of microbial communities. In this study, Baiyangdian Lake was selected as the study area. Sediment samples of 16 sites were collected in August 2018 and April 2019, respectively. Ultra-high performance liquid chromatography-mass spectrometry （HPLC-MS/MS） was used to determine the content of typical antibiotics-quinolones （QNs） in sediment. Through high-throughput sequencing technology, the structure and function of microbial communities was analyzed in the sediments to explore the spatiotemporal variation. Thereinto, redundancy analysis （RDA） was used to identify the key influencing factors of spatiotemporal variation of microbial communities. The results showed that： ① From August to April, the average ω（QNs） showed an increasing trend, and its mean value changed from 3.91 ng·g-1 to 6.34 ng·g-1, with significant seasonal differences in oxolinic acid （OXO） and total QNs content （P &lt; 0.05）. ② In terms of temporal variation, the dominant bacteria were Proteobacteria and Chloroflexi. The relative abundance of Proteobacteria showed a decreasing trend, whereas Chloroflexi showed an increasing trend； at the genus level, the dominant bacteria genera in August were norank_ f__Anaerolineaceae and Thiobacilus, and the dominant bacteria genera in April were Acinetobacter and norank_ f_Anaerolineaceae, and the dominant bacteria genera had significant seasonal differences （P &lt; 0.05）. ③ In terms of temporal variation, the index of Simpson, Chao, Ace, and OTU number all showed an increasing trend, and the seasonal differences were significant （P &lt; 0.05）. ④ In terms of spatial variation, there were no significant spatial differences among functional genes of COG. In terms of temporal variation, there were significant seasonal differences in functional genes of energy production and conversion, carbohydrate transport and metabolism, transcription, cell wall/membrane/envelope biogenesis, and signal transduction mechanisms （P &lt; 0.01 and P &lt; 0.05）. ⑤ Microbial community structure and functional genes were significantly correlated with QNs （P &lt; 0.01 and P &lt; 0.05）, and QNs were the main influencing factors. Therefore, QNs were the main factor affecting the changes in microbial community structure and functional genes in sediments of Baiyangdian Lake. Thus, comprehensive control of antibiotic pollution in sediments should be further strengthened.

Studies on Nigerian cashew nut shell liquid: Greening-up extraction process, chemical composition and cost effectiveness

Extractions of cashew nut shell liquid (CNSL) have always widely been carried out with non-environmentally friendly solvents. Hence, greener solvents are desired as alternative to currently used solvents. Herein, batch extractions of Nigerian CNSL with greener solvents: ethyl acetate and cyclohexane mixture in different ratios were investigated in hot and cold system and results compared with conventional solvent (petroleum ether). Compositional studies of the extracted CNSL and its isolated components were done with Fourier-Transformed Infrared (FTIR), Nuclear Magnetic Resonance (NMR) spectroscopy and Ultra High-Performance Liquid Chromatography-Mass Spectrometry (UHPLC-MS). Cost effectiveness analysis for the extraction solvents and methods was also investigated. It was revealed that ethyl acetate and ethyl acetate-cyclohexane mixture at cold and hot conditions recorded higher CNSL yields than previous reports even at a lower liquid-to-solid ratio. Chemical composition and component quantity extracted did not significantly change under hot or cold system. The present composition of the Nigerian CNSL contains unusually higher amount of anarcardic acid (79 %) and cardanol (14 %) than ever reported. Additionally, degree of unsaturation in the alkyl chain of the CNSL is in the order triene>monoene>diene in the anarcardic acid and cardanol components of the liquid. Cost effectiveness analysis at a kilogramme scale revealed that the order of greenness of extraction route based on solvent type, energy requirement and condition of extraction is ethyl acetate-solvent cold system> ethyl acetate-hot system> ethyl acetate-cyclohexane-system> cyclohexane system > pet. ether system.

Optimization of Phenolic-Enriched Extracts from Olive Leaves via Ball Milling-Assisted Extraction Using Response Surface Methodology.

This study aims to extract phenolic-enriched compounds, specifically oleuropein, luteoloside, and hydroxytyrosol, from olive leaves using ball milling-assisted extraction (BMAE). Response surface methodology (RSM) and the Box-Behnken design (BBD) were used to evaluate the effects of the temperature, solvent-to-solid ratio, and milling speed on extraction recovery. The contents of the extract were determined by ultra-high-performance liquid chromatography-mass spectrometry (UPLC-MS) and converted to recoveries to evaluate the extraction efficiency. The optimal extraction conditions for oleuropein, luteoloside, and hydroxytyrosol were identified. Oleuropein had a recovery of 79.0% ± 0.9% at a temperature of 56.4 °C, a solvent-to-solid ratio of 39.1 mL/g, and a milling speed of 429 rpm. Luteoloside's recovery was 74.6% ± 1.2% at 58.4 °C, 31.3 mL/g, and 328 rpm. Hydroxytyrosol achieved 43.1% ± 1.3% recovery at 51.5 °C, 32.7 mL/g, and 317 rpm. The reason for the high recoveries might be that high energy ball milling could reduce the sample size further, breaking down the cell walls of olive leaves, to enhance the mass transfer of these components from the cell to solvent. BMAE is displayed to be an efficient approach to extracting oleuropein, luteoloside, and hydroxytyrosol from olive leaves, which is easy to extend to industrial production.