As an essential amino acid, l-tryptophan is widely used in food, feed and medicine sectors. Nowadays, microbial l-tryptophan production suffers from low productivity and yield. Here we construct a chassis E. coli TRP3 producing 11.80 g/L l-tryptophan, which was generated by knocking out the l-tryptophan operon repressor protein (trpR) and the l-tryptophan attenuator (trpL), and introducing the feedback-resistant mutant aroG^fbr. On this basis, the l-tryptophan biosynthesis pathway was divided into three modules, including the central metabolic pathway module, the shikimic acid pathway to chorismate module and the chorismate to tryptophan module. Then we used promoter engineering approach to balance the three modules and obtained an engineered E. coli TRP9. After fed-batch cultures in a 5 L fermentor, tryptophan titer reached to 36.08 g/L, with a yield of 18.55%, which reached 81.7% of the maximum theoretical yield. The tryptophan producing strain with high yield laid a good foundation for large-scale production of tryptophan.
Escherichia coli/metabolism* ; Tryptophan ; Metabolic Engineering ; Bioreactors ; Metabolic Networks and Pathways

This study aims to explore the effect of tryptanthrin on potential metabolic biomarkers in the serum of mice with ulcerative colitis(UC) induced by dextran sulfate sodium(DSS) based on liquid chromatography-mass spectrometry(LC-MS) and predict the related metabolic pathways. C57BL/6 mice were randomly assigned into a tryptanthrin group, a sulfasalazine group, a control group, and a model group. The mouse model of UC was established by free drinking of 3% DSS solution for 11 days, and corresponding drugs were adminsitrated at the same time. The signs of mice were observed and the disease activity index(DAI) score was recorded from the first day. Colon tissue samples were collected after the experiment and observed by hematoxylin-eosin(HE) staining. The levels of interleukin-4(IL-4), interleukin-10(IL-10), tumor necrosis factor-α(TNF-α), interleukin-6(IL-6), and interleukin-8(IL-8) in the serum were measured by enzyme linked immunosorbent assay(ELISA). The serum samples were collected from 6 mice in each group for widely targeted metabolomics. The metabolic pathways were enriched by MetaboAnalyst 5.0. The results showed that compared with the model group, tryptanthrin treatment decreased the DAI score(P<0.05), alleviated the injury of the colon tissue and the infiltration of inflammatory cells, lowered the levels of proinflammatory cytokines, and elevated the levels of anti-inflammatory cytokines in the serum. The metabolomic analysis revealed 28 differential metabolites which were involved in 3 metabolic pathways including purine metabolism, arachidonic acid metabolism, and tryptophan metabolism. Tryptanthrin may restore the metabolism of the mice with UC induced by DSS to the normal level by regulating the purine metabolism, arachidonic acid metabolism, and tryptophan metabolism. This study employed metabolomics to analyze the mechanism of tryptanthrin in the treatment of UC, providing an experimental basis for the utilization and development of tryptanthrin.
Mice ; Animals ; Colitis, Ulcerative/drug therapy* ; Tryptophan ; Arachidonic Acid/metabolism* ; Mice, Inbred C57BL ; Colon ; Cytokines/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6/metabolism* ; Metabolomics ; Purines/therapeutic use* ; Dextran Sulfate/metabolism* ; Disease Models, Animal ; Colitis/chemically induced*

In recent years, reports of adverse reactions related to traditional Chinese medicine(TCM) have been on the rise, especially some traditionally considered "non-toxic" TCM(such as Dictamni Cortex). This has aroused the concern of scholars. This study aims to explore the metabolomic mechanism underlying the difference in liver injury induced by dictamnine between males and females through the experiment on 4-week-old mice. The results showed that the serum biochemical indexes of liver function and organ coefficients were significantly increased by dictamnine(P<0.05), and hepatic alveolar steatosis was mainly observed in female mice. However, no histopathological changes were observed in the male mice. Furthermore, a total of 48 differential metabolites(such as tryptophan, corticosterone, and indole) related to the difference in liver injury between males and females were screened out by untargeted metabolomics and multivariate statistical analysis. According to the receiver operating characteristic(ROC) curve, 14 metabolites were highly correlated with the difference. Finally, pathway enrichment analysis indicated that disorders of metabolic pathways, such as tryptophan metabolism, steroid hormone biosynthesis, and ferroptosis(linoleic acid metabolism and arachidonic acid metabolism), may be the potential mechanism of the difference. Liver injury induced by dictamnine is significantly different between males and females, which may be caused by the disorders of tryptophan metabolism, steroid hormone biosynthesis, and ferroptosis pathways.
Female ; Male ; Animals ; Mice ; Tryptophan ; Metabolomics ; Fatty Liver ; Steroids ; Hormones

This study aimed to investigate the mechanism of Psoraleae Fructus in improving the learning and memory ability of APP/PS1 mice by serum metabolomics, screen the differential metabolites of Psoraleae Fructus on APP/PS1 mice, and reveal its influence on the metabolic pathway of APP/PS1 mice. Thirty 3-month-old APP/PS1 mice were randomly divided into a model group and a Psoraleae Fructus extract group, and another 15 C57BL/6 mice of the same age were assigned to the blank group. The learning and memory ability of mice was evaluated by the Morris water maze and novel object recognition tests, and metabolomics was used to analyze the metabolites in mouse serum. The results of the Morris water maze test showed that Psoraleae Fructus shortened the escape latency of APP/PS1 mice(P<0.01), and increased the number of platform crossing and residence time in the target quadrant(P<0.01). The results of the novel object recognition test showed that Psoraleae Fructus could improve the novel object recognition index of APP/PS1 mice(P<0.01). Eighteen differential metabolites in serum were screened out by metabolomics, among which the levels of arachidonic acid, tryptophan, and glycerophospholipid decreased after drug administration, while the levels of glutamyltyrosine increased after drug administration. The metabolic pathways involved included arachidonic acid metabolism, glycerophospholipid metabolism, tryptophan metabolism, linoleic acid metabolism, α-linolenic acid metabolism, and glycerolipid metabolism. Therefore, Psoraleae Fructus can improve the learning and memory ability of APP/PS1 mice, and its mechanism may be related to the effects in promoting energy metabolism, reducing oxidative damage, protecting central nervous system, reducing neuroinflammation, and reducing Aβ deposition. This study is expected to provide references for Psoraleae Fructus in the treatment of Alzheimer's disease(AD) and further explain the mechanism of Psoraleae Fructus in the treatment of AD.
Mice ; Animals ; Amyloid beta-Protein Precursor/genetics* ; Mice, Transgenic ; Arachidonic Acid ; Tryptophan ; Mice, Inbred C57BL ; Alzheimer Disease/genetics* ; Maze Learning ; Glycerophospholipids ; Disease Models, Animal ; Amyloid beta-Peptides/metabolism*

This study aims to investigate the mechanism of Linderae Radix water extract(LRWE) in the prevention and treatment of diarrhea-predominant irritable bowel syndrome(IBS-D) based on serum metabolomics. Eighteen 2-week-old male SD rats were randomized into control, IBS-D model, and LRWE groups. The rats in other groups except the control group received gavage of senna concentrate combined with restraint stress for the modeling of IBS-D. The rats in the LRWE group were administrated with LRWE(5.4 g·kg~(-1)) by gavage, and those in the control and IBS-D model groups with an equal volume of distilled water for a total of 14 days. The visceral sensitivity was evaluated by the abdominal withdrawal reflex(AWR) score, and the degree of diarrhea was assessed by the fecal water content(FWC). The morphological changes of the colon and the morphology and number of goblet cells were observed by hematoxylin-eosin(HE) and periodic acid-schiff(PAS) staining, respectively. Ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS) was used for the screening of the potential biomarkers in the rat serum and their related metabolic pathways. The results showed that LRWE reduced the AWR score, decreased FWC, and alleviated visceral sensitivity and diarrhea symptoms in IBS-D rats. HE and PAS staining showed that LRWE mitigated low-grade intestinal inflammation and increased the number of mature secretory goblet cells in the colonic epithelium of IBS-D rats. A total of 25 potential biomarkers of LRWE in treating IBS-D were screened out in this study, which were mainly involved in riboflavin, tryptophan, glycine, serine and threonine metabolism, glyoxylate and dicarboxylate metabolism, and cysteine and methionine metabolism. The regulatory effects were the most significant on the riboflavin and tryptophan metabolism pathways. LRWE may alleviate the visceral hypersensitivity by promoting energy metabolism and amino acid metabolism, enhancing intestinal barrier function, and improving intestinal immune function in IBS-D rats.
Rats ; Male ; Animals ; Irritable Bowel Syndrome/metabolism* ; Water ; Chromatography, Liquid ; Tryptophan ; Rats, Sprague-Dawley ; Tandem Mass Spectrometry ; Diarrhea/drug therapy* ; Biomarkers ; Riboflavin

Sepsis is a life-threatening organ dysfunction caused by dysregulated host responses to infection. Despite significant advances in anti-infective, immunomodulatory, and organ function support technologies, the precise and targeted management of sepsis remains a challenge due to its high heterogeneity. Studies have identified disturbed tryptophan (TRP) metabolism as a common mechanism in multiple diseases, which is involved in both immune regulation and the development of multi-organ damages. The rise of research on intestinal microflora has further highlighted the critical role of microflora-regulated TRP metabolism in pathogen-host interactions and the "cross-talk" among multi-organs, making it a potential key target for precision medicine in sepsis. This article reviews TRP metabolism, the regulation of TRP metabolism by the intestinal microflora, and the characteristics of TRP metabolism in sepsis, providing clues for further clinical targeting of TRP metabolism for precision medicine in sepsis.
Humans ; Gastrointestinal Microbiome/physiology* ; Tryptophan/metabolism* ; Precision Medicine ; Sepsis

OBJECTIVE: To clarify the functional effects of differential expression of ring finger and tryptophan-aspartic acid 2 (RFWD2) on dendritic development and formation of dendritic spines in cerebral cortex neurons of mice. METHODS: Immunofluorescent staining was used to identify the location and global expression profile of RFWD2 in mouse brain and determine the co-localization of RFWD2 with the synaptic proteins in the cortical neurons. We also examined the effects of RFWD2 over-expression (RFWD2-Myc) and RFWD2 knockdown (RFWD2-shRNA) on dendritic development, dendritic spine formation and synaptic function in cultured cortical neurons. RESULTS: RFWD2 is highly expressed in the cerebral cortex and hippocampus of mice, and its expression level was positively correlated with the development of cerebral cortex neurons and dendrites. RFWD2 expression was detected on the presynaptic membrane and postsynaptic membrane of the neurons, and its expression levels were positively correlated with the length, number of branches and complexity of the dendrites. In cultured cortical neurons, RFWD2 overexpression significantly lowered the expressions of the synaptic proteins synaptophysin (P < 0.01) and postsynapic density protein 95 (P < 0.01), while RFWD2 knockdown significantly increased their expressions (both P < 0.05). Compared with the control and RFWD2-overexpressing cells, the neurons with RFWD2 knockdown showed significantly reduced number of dendritic spines (both P < 0.05). CONCLUSION RFWD2 can regulate the expression of the synaptic proteins, the development of the dendrites, the formation of the dendritic spines and synaptic function in mouse cerebral cortex neurons through ubiquitination of Pea3 family members and c-Jun, which may serve as potential treatment targets for neurological diseases.
Animals ; Aspartic Acid/metabolism* ; Cerebral Cortex ; Dendritic Spines/metabolism* ; Mice ; Neurons/metabolism* ; Synapses ; Tryptophan/metabolism*

To investigate the effect of Rehmanniae Radix on depression-like behavior and monoamine neurotransmitters of chronic unpredictable mild stress(CUMS) model rats. CUMS combined with isolated feeding was used to induce the depression model of rats. The depression-like behavior of rats was evaluated by sucrose preference test, open field test, and forced swim test. Hematoxylin-Eosin(HE) staining was used to investigate the pathological changes of neurons in the CA1 and CA3 area of hippocampus. Ultra performance liquid chromatography-tandem mass spectrometry(UPLC-MS) was used to detect the contents of 5-hydroxytryptamine(5-HT), 5-hydroxyindoleacetic acid(5-HIAA), dopamine(DA), 3,4-dihydroxyphenylacetic acid(DOPAC), homovanillic acid(HVA), norepinephrine(NE), and 3-methoxy-4-hydroxyphenyl glycol(MHPG) in rats. Western blot was used to detect the protein expressions of tryptophan hydroxylase 2(TPH2), serotonin transporter(SERT), and monoamine oxidase A(MAO-A) in the hippocampus of rats. Compared with the normal group, depressive-like behavior of rats was obvious in the model group. The arrangements of neurons in the CA1 and CA3 area of hippocampus were loose and disorderly. The levels of 5-HT, 5-HIAA, and 5-HT/5-HIAA in the hippocampal area were decreased(P<0.01). The protein expression of TPH2 was decreased(P<0.01), but those of SERT and MAO-A were increased(P<0.01). In the Rehmanniae Radix groups with 1.8 g·kg~(-1) and 7.2 g·kg~(-1), the depression-like behavior of CUMS rats and pathological changes of neurons in CA1, CA3 area of hippocampus were improved. The protein expression of TPH2(P<0.05, P<0.01) was increased, and those of SERT and MAO-A were down-regulated(P<0.05, P<0.01). The levels of 5-HT, 5-HIAA, and 5-HT/5-HIAA in hippocampus were increased(P<0.05, P<0.01). The changes in DA, DOPAC, HVA, DA/(DOPAC +HVA), NE, DHPG, and NE/DHPG were not statistically significant. The results suggested that Rehmanniae Radix improved depression-like behavior of CUMS rats, and the mechanism might be related to the regulation of synthesis, transportation, and metabolism of 5-HT neurotransmitter in the hippocampus.
3,4-Dihydroxyphenylacetic Acid/pharmacology* ; Animals ; Antidepressive Agents/therapeutic use* ; Chromatography, Liquid ; Depression/drug therapy* ; Disease Models, Animal ; Dopamine ; Eosine Yellowish-(YS)/pharmacology* ; Hematoxylin/pharmacology* ; Hippocampus/metabolism* ; Homovanillic Acid/pharmacology* ; Hydroxyindoleacetic Acid/metabolism* ; Methoxyhydroxyphenylglycol/pharmacology* ; Monoamine Oxidase/metabolism* ; Neurotransmitter Agents/metabolism* ; Norepinephrine/pharmacology* ; Plant Extracts ; Rats ; Rehmannia/chemistry* ; Serotonin/metabolism* ; Serotonin Plasma Membrane Transport Proteins/pharmacology* ; Stress, Psychological/metabolism* ; Tandem Mass Spectrometry ; Tryptophan Hydroxylase/metabolism*

Content of multiple components (neochlorogenic acid,L-tryptophan,vicenin-2,isoquercitrin,and astragalin) in Moringa oleifera leaves was determined by high-performance liquid chromatography (HPLC),and the absolute content-time curves were plotted.Based on Fick's law of diffusion and Higbie's penetration theory,the parameters of the equations were calculated,and the measured results were substituted into the mathematical model to fit the equations.The n and a obtained from the equations on the decocting time factor and the solvent volume were close to each other.The dynamic models of the five components are as follows:■.The variation of the content of multiple components in M.oleifera leaves with time and solvent volume was explored.It was found that the content of the components was the highest when the leaves were decocted for 30 min with solvent volume 12 folds of the medicinal material.The dissolution and destruction of components and the diffusion movement of components are the main causes of the content change of M.oleifera leaves at different time and with different solvent volumes.The R~2of the linear equations on the content and the equations on the decocting process (5-30min and solvent volume 12-20 folds of the medicinal materials) was≥0.999 8 and≥0.9,respectively.Thus,the content determination and the decocting kinetic model had high accuracy,which can reflect the change law of the content of key components in M.oleifera leaves during the decoction.This study is expected to serve as a reference for optimizing the decocting technology.
Kinetics ; Moringa oleifera/chemistry* ; Plant Leaves/chemistry* ; Solvents ; Tryptophan/analysis*

This study investigated the potential mechanism of Cordyceps militaris(CM) against non-small cell lung cancer(NSCLC) based on serum untargeted metabolomics. Specifically, Balb/c nude mice were used to generate the human lung cancer A549 xenograft mouse model. The tumor volume, tumor weight, and tumor inhibition rate in mice in the model, cisplatin, Cordyceps(low-, medium-, and high-dose), and CM(low-, medium-, and high-dose) groups were compared to evaluate the influence of CM on lung cancer. Gas chromatography-mass spectrometry(GC-MS) was used for the analysis of mouse serum, SIMCA 13.0 for the compa-rison of metabolic profiles, and MetaboAnalyst 5.0 for the analysis of metabolic pathways. According to the pharmacodynamic data, the tumor volume and tumor weight of mice in high-dose CM group and cisplatin group decreased as compared with those in the model group(P<0.05 or P<0.01). The results of serum metabolomics showed that the metabolic profiles of the model group were significantly different from those of the high-dose CM group, and the content of endogenous metabolites was adjusted to different degrees. A total of 42 differential metabolites and 7 differential metabolic pathways were identified. In conclusion, CM could significantly inhibit the tumor growth of lung cancer xenograft mice. The mechanism is the likelihood that it influences the aminoacyl-tRNA biosynthesis, the metabolism of D-glutamine and D-glutamate, metabolism of alanine, aspartate, and glutamate, metabolism of glyoxylate and dicarboxylic acid, biosynthesis of phenylalanine, tyrosine, and tryptophan, arginine biosynthesis as well as nitrogen metabolism. This study elucidated the underlying mechanism of CM against NSCLC from the point of metabolites. The results would lay a foundation for the anticancer research and clinical application of CM.
Alanine/metabolism* ; Animals ; Arginine/metabolism* ; Aspartic Acid ; Carcinoma, Non-Small-Cell Lung/drug therapy* ; Cisplatin/pharmacology* ; Cordyceps ; Glutamic Acid ; Glutamine ; Glyoxylates/metabolism* ; Humans ; Lung Neoplasms/drug therapy* ; Metabolomics/methods* ; Mice ; Mice, Nude ; Nitrogen/metabolism* ; Phenylalanine/metabolism* ; RNA, Transfer/metabolism* ; Tryptophan/metabolism* ; Tyrosine/metabolism*