Microbial Metabolomics: From Methods to Translational Applications.

Most microbe-associated infectious diseases severely affect human health. However, clinical diagnosis of pathogenic diseases remains challenging due to the lack of specific and highly reliable methods. To better understand the diagnosis, pathogenesis, and treatment of these diseases, systems biology-driven metabolomics goes beyond the annotated phenotype and better targets the functions than conventional approaches. As a novel strategy for analysis of metabolomes in microbes, microbial metabolomics has been recently used to study many diseases, such as obesity, urinary tract infection (UTI), and hepatitis C. In this chapter, we attempt to introduce various microbial metabolomics methods to better interpret the microbial metabolism underlying a diversity of infectious diseases and inspire scientists to pay more attention to microbial metabolomics, enabling broadly and efficiently its translational applications to infectious diseases, from molecular diagnosis to therapeutic discovery.

Functional metabolomics innovates therapeutic discovery of traditional Chinese medicine derived functional compounds.

Traditional Chinese medicines (TCMs) produce chemically diverse functional compounds that are importantly chemical resource for facilitating new drug discovery and development against a diversity of diseases. However, modern exploration of TCM derived functional compounds is significantly hindered by the inefficient elucidation of pharmacological functions over past decades, because conventional research methods are incapable of efficiently elucidating therapeutic potential of TCM conferred by multiple functional compounds. Functional metabolomics has the priority-capacity to characterize systems therapeutic actions of TCM by precisely capturing molecular interactions between disease response metabolite biomarkers (DRMB) and functional compounds (secondary metabolites), which underline pharmacological efficiency and associated therapeutic mechanisms. In this critical review, we innovatively summarize systems therapeutic feature of TCM derived functional compounds from a functional-metabolism perspective, then systems metabolic targets (SMT) identified by functional metabolomics method are strategically proposed to better understanding of therapeutic discovery of TCM derived functional compounds. In addition, we propose the perspective strategy as Spatial Temporal Operative Real Metabolomics (STORM) to considerably improve analytical capacity of functional metabolomics method by selectively incorporating the cutting edge technologies of mass spectrometry imaging, isotope-metabolic fluxomics, synthetic and biosynthetic chemistry, which could considerably enhance the precision and resolution of elucidating pharmacological efficiency and associated therapeutic mechanisms of TCM derived functional compounds. Collectively, such critical review is expected to provide novel perspective-strategy that could significantly improve modern exploration and exploitation of TCM derived functional compounds that further promote new drug discovery and development against the complex diseases.

Mass spectrometry based targeted metabolomics precisely characterized new functional metabolites that regulate biofilm formation in Escherichia coli.

Biofilms are broadly formed by diverse microorganisms under stressful environments that are basically surrounded by an EPS matrix, which enable bacterial cells to confer the resistance to the biocides, antibiotics and other invasions. Yet, biofilms cause harmful impacts in various fields, including clinical infections, food contaminations and environmental pollution. However, the mechanism of biofilm formation remains incompletely elucidated, and currently, we lack an efficient strategy to tackle these tough problems by eradicating biofilms. In the present study, we sought to decipher the mechanism of biofilm formation in Escherichia coli from metabolic perspective. By exposing bacterial cells to various concentrations of iron, we found that iron can regulate biofilm formation, and the phenotypic changes were obviously dependent on iron concentration. A functional metabolome assay was further implemented to investigate the regulatory mechanism of iron on biofilm formation; we verified that siderophores mostly account for the transportation of iron into bacterial cells. Then, the bioavailable iron was recruited by bacterial cells to direct the levels of five functional metabolites (l-tryptophan, 5'-MTA, spermidine, CMP and L-leucine), which were identified as new effectors that directly regulate biofilm formation. Taken together, this study is the first to identify five functional metabolites to efficiently regulate biofilm formation, which can be targeted to tackle the harmful impacts associated with biofilm formation in different niches.

Berberine improves colitis by triggering AhR activation by microbial tryptophan catabolites.

Inflammatory bowel diseases (IBD) are kind of recurrent inflammatory issues that occur in the gastrointestinal tract, and currently clinical treatment is still unideal due to the complex pathogenesis of IBD. Basically, gut barrier dysfunction is triggered by gut microbiota dysbiosis that is closely associated with the development of IBD, we thus investigated the therapeutic capacity of berberine (BBR) to improve the dysregulated gut microbiota, against IBD in rats, using a combinational strategy of targeted metabolomics and 16 s rDNA amplicon sequencing technology. Expectedly, our data revealed that BBR administration could greatly improve the pathological phenotype, gut barrier disruption, and the colon inflammation in rats with dextran sulfate sodium (DSS)-induced colitis. In addition, 16S rDNA-based microbiota analysis demonstrated that BBR could alleviate gut dysbiosis in rats. Furthermore, our targeted metabolomics analysis illustrated that the levels of microbial tryptophan catabolites in the gastrointestinal tract were significantly changed during the development of the colitis in rats, and BBR treatment can significantly restore such changes of the tryptophan catabolites accordingly. At last, our in vitro mechanism exploration was implemented with a Caco-2 cell monolayer model, which verified that the modulation of the dysregulated gut microbiota to change microbial metabolites coordinated the improvement effect of BBR on gut barrier disruption in the colitis, and we also confirmed that the activation of AhR induced by microbial metabolites is indispensable to the improvement of gut barrier disruption by BBR. Collectively, BBR has the capacity to treat DSS-induced colitis in rats through the regulation of gut microbiota associated tryptophan metabolite to activate AhR, which can greatly improve the disrupted gut barrier function. Importantly, our finding elucidated a novel mechanism of BBR to improve gut barrier function, which holds the expected capacity to promote the BBR derived drug discovery and development against the colitis in clinic setting.

Cell Metabolomics Reveals Berberine-Inhibited Pancreatic Cancer Cell Viability and Metastasis by Regulating Citrate Metabolism.

Pancreatic cancer (PC) is becoming one of the deadliest cancers, with mortality among the highest worldwide because of its pathogenic latency and the lack of efficient drugs in the clinic. Considering that cancer cells undergo proliferation and differentiation at substantial metabolic costs, as indicated by dysregulated glycolysis and an abnormal TCA cycle induced by mitochondrial damage, we investigated the therapeutic capacity of berberine (BBR) in pancreatic cancer using a cell metabolomics method. A phenotypic assay revealed the significant inhibitory role of BBR in PC cell viability and metastasis. In addition, a precision-targeted metabolome assay showed that BBR profoundly dysregulated the energy metabolism of PC cells, and phenotypic observations based on imaging indicated that PC cell mitochondria were markedly damaged after BBR treatment. Notably, citrate metabolism and transportation in cell mitochondria were significantly influenced by BBR, which led to the blocked biosynthesis of the defined fatty acids (FAs) through the regulation of ACLY, ACO1, and SLC25A1. Therefore, the regulatory effects of FAs on PC cell proliferation and metastasis may be regulated by BBR through targeting citrate metabolism. Collectively, our in vitro data preliminarily reveals the therapeutic potential of BBR against pancreatic cancer by targeting citrate metabolism, citrate might be a new target for drug development and the treatment against PC, but further experimental verification will be required subsequently. Moreover, our study demonstrated that the cell metabolomics method pertains to the capacity to rapidly explore biochemical functions of natural products.

Metabolomics identified new biomarkers for the precise diagnosis of pancreatic cancer and associated tissue metastasis.

Pancreatic cancer (PC) is one of the most aggressive malignancies with high mortality due to a complex and latent pathogenesis leading to the severe lack of early diagnosis methods. To improve clinical diagnosis and enhance therapeutic outcome, we employed the newly developed precision-targeted metabolomics method to identify and validate metabolite biomarkers from the plasma samples of patients with pancreatic cancer that can sensitively and efficiently diagnose the onsite progression of the disease. Many differential metabolites have the capacity to markedly distinguish patients with pancreatic cancer (n = 60) from healthy controls (n = 60). To further enhance the specificity and selectivity of metabolite biomarkers, a dozen tumor tissues from PC patients and paired normal tissues were used to clinically validate the biomarker performance. We eventually verified five new metabolite biomarkers in plasma (creatine, inosine, beta-sitosterol, sphinganine and glycocholic acid), which can be used to readily diagnose pancreatic cancer in a clinical setting. Excitingly, we proposed a panel biomarker by integrating these five individual metabolites into one pattern, demonstrating much higher accuracy and specificity to precisely diagnose pancreatic cancer than conventional biomarkers (CA125, CA19-9, CA242 and CEA); moreover, this plasma panel biomarker used for PC diagnosis is also quite convenient to implement in clinical practice. Using the same metabolomics method, we characterized succinic acid and gluconic acid as having a great capability to monitor the progression and metastasis of pancreatic cancer at different stages. Taken together, this metabolomics method was used to identify and validate metabolite biomarkers that can precisely and sensitively diagnose the onsite progression and metastasis of pancreatic cancer in a clinical setting. Furthermore, such effort should leave clinicians with the correct time frame to facilitate early and efficient therapeutic interventions, which could largely improve the five-year survival rate of PC patients by significantly lowering clinical mortality.

Omics strategies decipher therapeutic discoveries of traditional Chinese medicine against different diseases at multiple layers molecular-level.

Traditional Chinese medicine (TCM) has been broadly used for the personalized treatment of many diseases in China for thousands of years. In the past century, TCM was also introduced to other Asian countries and even the Western world. Increasing evidence has shown that TCM has the capacity to treat numerous complex diseases in the clinic, such as cardiovascular diseases (CVDs), infectious diseases, metabolic diseases, and neurodegenerative diseases. However, the earlier lack of analytical strategies to annotate the chemical complexity has severely impeded the modern study and translational application of TCM. This critical review aims to explore and exploit applications of systems biology-driven omics methods in TCM against a diversity of diseases, toward the specific use of TCM to treat patients with different diseases. Such effort shall enhance the applicability of systems biology-driven omics strategies in deciphering the mechanisms by which TCM treats different diseases and may lead to the discovery of new therapeutic directions. In addition, we proposed the possible strategies to innovate the applicable pattern of omics technologies in TCM niches, such as precision-modification metabolomics and chinmedomics methods, allowing to unveil the complexity of TCM, which must enable TCM to serve better for the population-health. Taken together, this review eventually shall highlight the core value of omics technologies in innovating TCM to combat the diseases in a new horizon.

Mass spectrometry and associated technologies delineate the advantageously biomedical capacity of siderophores in different pathogenic contexts.

Siderophores are chemically diverse small molecules produced by microorganisms for chelation of irons to maintain their survival and govern some important biological functions, especially those cause that infections in hosts. Still, siderophores can offer new insight into a better understanding of the diagnosis and treatments of infectious diseases from the siderophore biosynthesis and regulation perspective. Thus, this review aims to summarize the biomedical value and applicability of siderophores in pathogenic contexts by briefly reviewing mass spectrometry (MS)-based chemical biology and translational applications that involve diagnosis, pathogenesis, and therapeutic discovery for a variety of infectious conditions caused by different pathogens. We highlight the advantages and disadvantages of siderophore discovery and applications in pathogenic contexts. Finally, we propose a panel of new and promising strategy as precision-modification metabolomics method, to rapidly advance the discovery of and translational innovations pertaining to these value compounds in broad biomedical niches. © 2018 Wiley Periodicals, Inc. Mass Spec Rev XX:XX-XX, 2018.

Acylhydrazone bond dynamic covalent polymer gel monolithic column online coupling to high-performance liquid chromatography for analysis of sulfonamides and fluorescent whitening agents in food.

A new dynamic covalent polymer (DCP) gel was well designed and constructed based on imine chemistry. Polycondensation of 4,4'-biphenyldicarboxaldehyde and 1,3,5-benzenetricarbohydrazide via Schiff-base reaction resulted in an acylhydrazone bond gel (AB-gel) DCP. AB-gel DCP had three-dimensional network of interconnected nanoparticles with hierarchically porous structure. AB-gel DCP was successfully fabricated as a monolithic column by an in-situ chemical bonding method for online enrichment and separation purpose with excellent permeability. AB-gel DCP based monolithic column showed remarkable adsorption affinity towards target analytes including sulfonamides (SAs) and fluorescent whitening agents (FWAs) due to its strong π-π affinity, hydrophobic effect and hydrogen bonding interaction. Then, AB-gel DCP based monolithic column was applied for online separation and analysis of trace SAs and FWAs in food samples coupled with high-performance liquid chromatography (HPLC). Sulfathiazole (ST) and sulfadimidine (SM2) in one positive weever sample were actually found and determined with concentrations of 273.8 and 286.3µg/kg, respectively. 2,5-Bis(5-tert-butyl-2-benzoxazolyl) thiophene (FWA184) was actually quantified in one tea infusion sample with the concentration of 268.5ng/L. The spiked experiments suggested the good recoveries in range of 74.5-110% for SAs in weever and shrimp samples with relative standard deviations (RSDs) less than 9.7% and in range of 74.0-113% for FWAs in milk and tea infusion samples with RSDs less than 9.0%. AB-gel DCP monolithic column was proved to be a promising sample preparation medium for online separation and analysis of trace analytes in food samples with complex matrices.

In-tube solid-phase microextraction based on NH2-MIL-53(Al)-polymer monolithic column for online coupling with high-performance liquid chromatography for directly sensitive analysis of estrogens in human urine.

In this work, a novel NH2-MIL-53(Al) incorporated poly(styrene-divinylbenzene-methacrylic acid) (poly(St-DVB-MAA)) monolith was prepared via chemical fabrication. Moreover, it has been efficiently applied to the in-tube solid-phase microextraction (SPME) for online coupling with high-performance liquid chromatography (HPLC) to the direct determination of five estrogens in human urine samples. The NH2-MIL-53(Al)-polymer monolith was suitable for in-tube SPME owing to its good permeability, high extraction efficiency, chemical stability, good reproducibility and long lifetime. The extraction conditions including extraction solvent, pH of sample solution, flow rate of extraction and desorption, and desorption volume were investigated. Under the optimum conditions, the enrichment factors were 180-304 and saturated amounts of extraction were 2326-21393 pmol for estriol, 17ß-estradiol, estrone, ethinyl estradiol and progesterone, respectively. The adsorption mechanism was also explored which contributed to its strong extraction to target compounds. The proposed method had low limit of detection (2.0-40ng/L) and good linearity (with R2 between 0.9908 and 0.9978). Four endogenous estrogens were detected in urine samples and the recoveries of all five analytes were ranged from 75.1-120% with relative standard deviations (RSDs) less than 8.7%. The results showed that the proposed online SPME-HPLC method based on NH2-MIL-53(Al)-polymer monolithic column was highly sensitive for directly monitoring trace amount of estrogens in human urine sample.

[Progress in sample preparation and analytical methods for trace polar small molecules in complex samples].

Small polar molecules such as nucleosides, amines, amino acids are important analytes in biological, food, environmental, and other fields. It is necessary to develop efficient sample preparation and sensitive analytical methods for rapid analysis of these polar small molecules in complex matrices. Some typical materials in sample preparation, including silica, polymer, carbon, boric acid and so on, are introduced in this paper. Meanwhile, the applications and developments of analytical methods of polar small molecules, such as reversed-phase liquid chromatography, hydrophilic interaction chromatography, etc., are also reviewed.