Indole Concentrations Research Articles

Mixture Effects of Tryptophan Intestinal Microbial Metabolites on Aryl Hydrocarbon Receptor Activity.

Aryl hydrocarbon receptor (AHR) plays pivotal roles in intestinal physiology and pathophysiology. Intestinal AHR is activated by numerous dietary, endogenous, and microbial ligands. Whereas the effects of individual compounds on AHR are mostly known, the effects of real physiological mixtures occurring in the intestine have not been studied. Using reporter gene assays and RT-PCR, we evaluated the combinatorial effects (3520 combinations) of 11 microbial catabolites of tryptophan (MICTs) on AHR. We robustly (n = 30) determined the potencies and relative efficacies of single MICTs. Synergistic effects of MICT binary mixtures were observed between low- or medium-efficacy agonists, in particular for combinations of indole-3-propionate and indole-3-lactate. Combinations comprising highly efficacious agonists such as indole-3-pyruvate displayed rather antagonist effects, caused by saturation of the assay response. These synergistic effects were confirmed by RT-PCR as CYP1A1 mRNA expression. We also tested mimic multicomponent and binary mixtures of MICTs, prepared based on the metabolomic analyses of human feces and colonoscopy aspirates, respectively. In this case, AHR responsiveness did not correlate with type of diet or health status, and the indole concentrations in the mixtures were determinative of gross AHR activity. Future systematic research on the synergistic activation of AHR by microbial metabolites and other ligands is needed.

Purification of Indole Contained in Wash Oil by Combination of Extraction and Crystallization (Part 1: Recovery and Concentration of Indole Contained in Wash Oil by Solvent Extraction).

For the purpose of determining the high-purity purification of indole (IN) contained in wash oil in concentrations of 5.75%, this study, first of all, investigated the concentration of IN contained in wash oil using a combination of methanol extraction to recover IN in the wash oil, and n-hexane re-extraction to concentrate IN present in the extract phase, recovered through methanol extraction. In order to examine the effect of each operation on the recovery and concentration of IN contained in the wash oil, batch 1-stage and batch co-current 5-stage distribution equilibrium was performed simultaneously. As 5-stage equilibrium extraction and re-extraction progressed, the recovery rate of IN decreased to about 79.1%, while IN composition in the raffinate phase recovered by re-extraction was highly concentrated to about 73.3%. From the high recovery rate and composition of IN obtained through this study, we confirmed that the combination examined by this study was one of the very useful combinations for the concentration of IN present in the wash oil. Furthermore, we reviewed the recovery and concentration process of IN contained in the wash oil using the experiment results of the extraction and the re-extraction obtained from this study.

Probiotics or synbiotics addition to sows' diets alters colonic microbiome composition and metabolome profiles of offspring pigs.

Little information exists about the effects of maternal probiotics and synbiotics addition on the gut microbiome and metabolome of offspring. The present study evaluated the effects of probiotics or synbiotics addition to sows’ diets on colonic microbiota and their metabolites in offspring using 16S rRNA gene sequencing and metabolome strategy. A total of 64 pregnant Bama mini-pigs were randomly divided into control, antibiotic, probiotics, and synbiotics groups and fed the corresponding experimental diets during pregnancy and lactation. After weaning, two piglets per litter and eight piglets per group were selected and fed a basal diet. The β-diversity analysis showed that the colonic microbiota of offspring had a clear distinction among the four groups at 65 days of age. Maternal probiotics addition increased the Actinobacteria abundance at 65 days of age and Tenericutes and Firmicutes abundances at 95 days of age of offspring compared with the other three groups, whereas maternal antibiotic addition increased Spirochaetes and Proteobacteria abundances at 95 days of age of offspring compared with the other three groups. Metabolomic analysis showed that colonic metabolites were different between the groups, regardless of the days of age. Furthermore, both PICRUSt2 and enrichment analysis of metabolic pathways showed that maternal probiotics and synbiotics addition affected metabolism of carbohydrate, amino acid, cofactors and vitamins in the colonic microbiota. Compared with the control group, the colonic concentration of indole decreased and skatole increased in the probiotics group, whereas indole increased and skatole decreased in the synbiotics group. Maternal probiotics addition increased the colonic concentrations of acetate and butyrate at 65 and 125 days of age, whereas probiotics and synbiotics addition decreased short-chain fatty acids concentrations at 95 days of age. In addition, the colonic concentrations of putrescine, cadaverine, 1,7-heptanediamine, and spermidine were increased in the antibiotic, probiotics, and synbiotics groups compared with the control group at 95 days of age. The correlation analysis showed that Gemmiger, Roseburia, and Faecalibacterium abundances were positively correlated with acetate, propionate, and butyrate concentrations; Gemmiger, Blautia, and Faecalibacterium were positively correlated with putrescine and spermidine; and Faecalibacterium, Blautia, Clostridium, and Streptococcus were positively correlated with (R)-3-hydroxybutyric acid. Collectively, these findings suggest that probiotics and synbiotics addition to sows’ diets exerts effects on offspring pigs by altering gut microbiota composition and their metabolites. The potential beneficial effect on gut health is discussed.

A Selective and Sensitive LC-MS/MS Method for Quantitation of Indole in Mouse Serum and Tissues.

Indole is an endogenous substance currently being evaluated as a biomarker for ulcerative colitis, irritable bowel syndrome, Crohn’s disease and non-alcoholic fatty liver disease. A novel, selective, and sensitive method using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) was developed for quantitation of indole concentrations in mouse plasma and tissues. Samples were prepared by protein precipitation using ice-cold acetonitrile (ACN) followed by injecting the extracted analyte to LC-MS/MS system. Indole was separated using Synergi Fusion C18 (4 µm, 250 × 2.0 mm) column with mobile phase 0.1% aqueous formic acid (A) and methanol (B) using gradient flow with run time 12 min. The mass spectrometer was operated in atmospheric pressure chemical ionization (APCI) positive mode at unit resolution in multiple reaction monitoring (MRM) mode, using precursor ion > product ion combinations of 118.1 > 91.1 m/z for indole and 124.15 > 96.1 m/z for internal standard (IS) indole d7. The MS/MS response was linear over the range of indole concentrations (1–500 ng/mL). The validated method was applied for quantitation of indole concentrations range in mouse lungs (4.3–69.4 ng/g), serum (0.8–38.7 ng/mL) and cecum (1043.8–12,124.4 ng/g). This method would help investigate the role of indole as a biomarker and understand its implications in different disease states.

Is a short concentrate-finishing period in lambs raised on alfalfa pasture effective at reducing fat indoles contents and lightening meat colour?

Pasture-feeding in lambs has been associated with dark-coloured meat and high meat contents of volatile indoles (skatole and indole), which may be responsible for sensory defects. The risk of off-flavours is even higher when lambs graze alfalfa, because it is particularly rich in rapidly degradable proteins. Here, we investigated whether a short concentrate stall-finishing period in lambs that previously grazed alfalfa influences meat sensory quality traits. We compared three feeding treatments, using three groups of 10 male Romane lambs: grazing alfalfa (A), stall-feeding with concentrate and straw (S), and stall-finishing with concentrate and straw for 21 days after previously grazing alfalfa (AS). During stall-feeding, the concentrate was given at a level adjusted to achieve a similar growth pattern in all treatment groups, and the length of the trough was sufficient for all lambs to have access to the feeds at the same time. Lamb live weight and age at slaughter, and carcass conformation and fatness were similar for all treatment groups. Perirenal fat skatole content was lower in AS and S lambs than in A lambs (P < 0.001 and P < 0.05, respectively), and not significantly different between AS and S lambs. Perirenal fat indole and dorsal fat skatole concentrations were lower in AS lambs than in A lambs (P < 0.05 and P < 0.005, respectively), the other pairwise comparisons being not significant. There was no treatment effect on dorsal fat indole content. Longissimus et lumborum muscle colour coordinates differed between pasture-fed and stall-fed lambs, with a lower lightness (P < 0.005) and greater redness and hue angle (P < 0.001 and P < 0.01, respectively) in pasture-fed lambs, but muscle colour coordinates did not differ between A and AS lambs. A 21-d concentrate-based stall-finishing period in lambs previously raised on alfalfa pasture therefore markedly reduced fat indoles concentrations compared to lambs grazed on alfalfa until slaughter but did not change meat colour.

Regulatory effect of polyamines and indole on expression of stress adaptation genes in &lt;i&gt; Escherichia coli &lt;/i&gt;

Background. Indole and polyamines are involved in the regulation of physiological processes in bacteria associated with adaptation to stress, biofilm formation, antibiotic tolerance, and bacterial persistence. However, the molecular targets and mechanisms of action of these metabolites are still poorly understood. In this work, we studied the effect of polyamines and indole on the expression of such genes as: rpoS, relA, and spoT, encoding regulators of the general stress responses and starvation; hns and stpA, encoding global regulators of gene expression; rmf, yqjD, hpf, raiA, rsfS, sra, ettA, encoding ribosome hibernation factors.The aim. To study the regulatory effects of polyamines and indole on the expression of these genes, which are responsible for the adaptation of Escherichia coli to stress.Materials and methods. We used strains of E. coli in this study. The amount of polyamines was studied by thin layer chromatography. The indole concentration was determined by high performance liquid chromatography. Gene expression was studied using real-time RT-PCR.Results. The addition of polyamines putrescine, cadaverine and spermidine to the medium stimulated the expression of all the studied genes. The maximal stimulation was observed at the stationary phase mostly. Putrescine and spermidine had the most significant effect. At 24 h of cultivation, an equimolar conversion of exogenous tryptophan into indole was showed. At this time, the expression of two genes – rmf and raiA – increased.Conclusions. We have shown that polyamines upregulate the expression of all the studied genes at the transcriptional level. The stimulating effect is specific for the phase of the batch culture and the type of polyamine. Indole has a positive effect on the expression of the rmf and raiA genes.

Concentrations of Boar Taint Compounds Are Weakly Associated with Sexual Behavior of Young Boars.

Simple SummaryBoar taint is an off-odor and off-flavor affecting pork meat caused by the accumulation of three main compounds in the adipose tissue of uncastrated male pigs: androstenone, skatole, and indole. Boar taint occurrence is associated with sexual maturity onset; thus, strategies for reducing boar taint compound concentrations might lead to impaired male sexual behavior. This study investigated the relationships between boar taint compound concentrations and sexual behavior traits (sexual arousal, salivation, mounting performance, interest in the dummy sow, penis unsheathing, and overall libido) scored during training with the dummy sow in 391 young boars. Overall, our results suggest that a weak relationship between boar taint compounds and sexual behavior traits exists. The maximization of libido performances was observed at high levels of androstenone, intermediate or low levels of skatole, and intermediate to high levels of indole, but the relationship between BT compounds and sexual behavior appears to be weak.Strategies to control boar taint (BT) in meat relies on the reduction of skatole, indole, and androstenone concentration. This might have unfavorable effects on the libido of breeding boars. The association between BT compound concentration in backfat and libido was investigated in 391 commercial breeding boars. Six sexual behavior traits (SBT; sexual arousal, salivation, mounting performance, interest in the dummy sow, penis unsheathing, and overall libido score) were scored during the training of the boars with the dummy sow. Variation in SBT was analyzed by proportional-odds cumulative logistic models. Overall, indole, skatole, and androstenone concentrations were weakly associated with libido. Farm of origin, age at training or body weight, and BT compound levels were poor predictors of boar performance (the area under the ROC curve ranged from 0.60 to 0.69). This indicates that BT compound concentrations were weakly associated with libido, even though the probability of observing good SBT scores increased with high levels of androstenone, intermediate or low levels of skatole, and intermediate to high levels of indole. Hence, practices aiming at reducing androstenone, and controlling the concentrations of skatole and indole to intermediate levels are not expected to impair the libido of young boars.

Association of Microbial Tryptophan Metabolites and Indoleamine-2,3-Dioxygenase with Immune Activity in Healthy Adults

ObjectivesTryptophan (Trp) metabolites from intestinal bacteria, including indole, indole acetic acid (IAA) and indole propionic acid (IPA), and from the mammalian indoleamine 2,3-dioxygenase (IDO) pathway, including kynurenine (Kyn), can regulate immune activation. IDO activity in vivo is measured by the plasma Kyn/Trp ratio. In this study we hypothesized that (1) the plasma Kyn/Trp ratio, (2) plasma indole, IPA, and IAA concentrations, and (3) intestinal bacteria associated with indole metabolism, would all be associated with inflammation and immune activation markers in a study population of healthy adults.MethodsA group of 88 immune markers was assessed using plasma, flow cytometric analysis of whole blood and of peripheral blood mononuclear cells (PBMC), and ex vivo culture of PBMC, in 362 healthy, fasting adults. Plasma Trp metabolites were also measured. Bacterial taxa from stool were identified by 16S rRNA gene analysis and those associated with plasma Trp metabolites were identified. Multiple linear regression analysis was used to identify statistically significant associations (with p < .05 after adjustment for multiple comparisons) of these metabolites and taxa with immune markers.ResultsSeveral robust positive associations were found between Kyn/Trp and markers of inflammation including neopterin, IP-10, TNF-α, C-reactive protein, and the regulatory cytokine IL-10. A significant positive association was found between the sum of indole and IAA and natural killer T-cells. Three taxa associated with bacterial Trp metabolites were associated with elevated markers of immune activation: the family Lachnospiraceae with higher lymphocyte counts, the genus Dorea with higher production of the type 1 cytokine IFN-γ by T-cells in PBMC culture, the genus Ruminococcus with higher production of the inflammatory cytokine IL-6 in PBMC cultures stimulated with bacterial lipopolysaccharide.ConclusionsIDO activity, bacterial Trp metabolites and indole-associated commensal bacteria were all associated with markers of systemic immune activation, particularly involving inflammatory cytokines, the acute phase response and type 1 innate and adaptive immunity. These findings suggest sub-clinical levels of systemic and/or intestinal immune activity that might be precursors to adverse health outcomes.Funding SourcesUSDA/ARS 2032-51,530-026-00D.

Wheatgerm Supplementation Reduces Gut Inflammation and Epithelial Barrier Dysfunction in IL-10 KO Mice Fed Atherogenic Diet

Abstract Objectives Wheat germ (WG) contains many bioactive compounds with the potential to maintain an anti-inflammatory gut environment. This study investigated the effects of WG supplementation on gut inflammation and integrity in high-fat fed interleukin (IL)-10 KO mice. Methods Eight-wk-old female B6.129P2-Il10tm1Cgn/J (IL-10KO) and C57BL/6 (WT) mice (n = 10/group) were randomly assigned to diets: WT fed a control diet (WTCO; AIN93-M) and IL-10 KO mice fed control (KOCO), high-fat with high-cholesterol (HFHC; 45% fat kcal, 1% cholesterol), or HFHC + 10% WG (HFWG) for 3 m. Disease activity indices (fecal blood, ruffled fur, stool softness, and rectal prolapse) were monitored twice a week. Fecal indole and short chain fatty acids (SCFAs) concentration were assessed at the beginning and end of study. Proinflammatory cytokines were assessed in the serum and ileum. Ileal and colonic protein expression of transcription factors (STAT3, p-STAT3, PPARg, FoxP3, and AhR), tight junction proteins (ZO-1, occludin), and tryptophan catabolizing enzyme (IDO-1) were assessed by immunoblotting. Relative ileal and colonic gene expression of IL-22 and antimicrobial peptides (Reg3b and Reg3g) were assessed using qRT-PCR. P &amp;lt; 0.05 was considered statistically significant. Results WG increased (P = 0.003) colon length compared to the HFHC group. Weight loss (12.2% in HFHC vs WTCO) was not prevented by WG, but disease activity indices were significantly reduced in the WG vs HFHC group. WG also increased fecal indole, total SCFAs and acetate accompanied by an increase in colonic protein expression of PPARg (P &amp;lt; 0.0001) and FoxP3 (P = 0.001). Ileal STAT3 phosphorylation was reduced (P = 0.0076) due to WG supplementation. An increased colon and ileal protein expression of IDO-1 in the HFHS group was reduced by WG, while also increasing the expression of AhR, ZO-1, and occludin. The relative gene expression of the antimicrobial peptides (Reg3b and Reg3g) was increased (P &amp;lt; 0.05) while serum and ileal tissue concentration of the proinflammatory cytokine, IL-17 was reduced (P = 0.0165 and p = 0.0248 respectively) by WG. Conclusions WG modulated changes that are associated with HF-feeding in IL-10 KO mice, and might be a promising regimen for ameliorating the effects of gut inflammation. Funding Sources Oklahoma Agriculture Experiment Station, Jim and Lynn Williams Professorship, and Barbara K. Pass Grant.

Translational Detection of Indole by Complementary Cell-free Protein Synthesis Assay.

The information encoded in a single copy of DNA is processed into a plethora of protein molecules via the cascade of transcription and translation. Thus, the molecular process of gene expression can be considered an efficient biological amplifier from the viewpoint of synthetic biology. Cell-free protein synthesis (CFPS) enables the implementation of this amplification module for in vitro analysis of important biomolecules and avoids many of the problems associated with whole cell-based approaches. Here, we developed a method to analyze indole by using a combination of enzymatic conversion of indole and amino acid-dependent CFPS. In this method, indole molecules in the assay sample are used to generate tryptophan, which is incorporated into signal-generating proteins in the subsequent cell-free synthesis reaction. The activity of cell-free synthesized proteins was successfully used to estimate the indole concentration in the assay sample. In principle, the developed method could be extended to analyses of other important bioactive compounds.

MO436: The Effect of Protein-Restriction on the Tubular Handling of the Uraemic Retention Solute Indoxyl Sulfate

Abstract BACKGROUND AND AIMS Chronic kidney disease (CKD) patients present an increased plasma level of protein-bound solutes known as uraemic toxins. Their accumulation is associated with, amongst others, cardiovascular events. The gut microbiome plays an important role in the generation of part of the uraemic toxins. It has been postulated that proximal tubule cells in kidneys are able to sense augmented levels of gut microbial metabolites via receptors and signalling pathways [1]. According to the so-called remote sensing hypothesis, kidney tubular cells respond to uraemic toxins plasma variations modulating the activity of membrane transporters involved in their excretion. Indoxyl sulfate (IS) originates from the microbial fermentation of tryptophan, a diet-derived amino acid. Nonetheless, few studies have investigated whether gut generation, absorption and plasma retention of indolic uraemic toxins and kidney cells’ secretory mechanisms are affected by the protein content in the diet. METHOD A total of 18 Sprague–Dawley male rats (Janvier, Le Genest- St Isle, France) 7–8 weeks old (weighing 270–388 g) were induced with CKD utilizing 5/6 nephrectomy and were randomly assigned to a low protein (LP) (n = 10) or a high protein (HP) (n = 8) diet. A sham-operated control group for each diet was used (n = 7 and n = 8 respectively). 24 h-urine was collected after 7 weeks, and euthanasia was carried out after 8 weeks from the induction of the disease (Figure 1). Blood and colon samples were collected. Diffusion chambers were used to assess colon permeability to indole over time (t = 0, 60, 90, 120 min). Indole concentration was quantified by Kovacs assay. Blood creatinine and urea were determined with standard laboratory techniques. Total plasmatic and urinary IS concentrations were measured using LC-MS/MS. The fractional excretion (FE) i.e. the percentage of IS excreted relatively to the kidney filtered load, was calculated to assess IS remote sensing. RESULTS Plasmatic levels of IS were significantly higher (P &amp;lt;0.001) in CKD rats compared to sham rats. However, CKD rats on an HP diet showed no difference in plasma IS (P = 0.63) compared to rats on an LP diet. Conversely, 24-h urinary IS was significantly increased in CKD rats on a HP diet (P &amp;lt;0.001). The FE of IS was significantly higher (P = 0.005) in CKD rats on an HP diet (Figure 2) and correlated with 24 h-urinary IS (Spearman r = 0.51, P -value = 0.03) and with protein intake (Spearman r = 0.52, P-value = 0.03) in CKD rats. Therefore, CKD rodent models’ results suggest higher absorption and/or production of indole at the intestinal level when an HP is administered to CKD rats. Plus, they provide additional proof of the remote sensing theory of indole metabolites. The permeability of colon to indole evaluated ex vivo with the diffusion chamber technique, showed an increased apparent apical-to-basolateral transport of indole (P = 0.049) in CKD rats on an HP diet compared to that of sham rats on the same diet, again suggesting modulatory mechanisms affecting indole metabolites transporters uptake. CONCLUSION An HP diet in 5/6 nephrectomized CKD rats leads to increased production of indole in the colon and increased fractional excretion of IS. These results provide an additional proof of mechanisms of remote sensing and signalling of indole-derived uraemic toxins. To conclude, the adoption of a low protein diet remains recommended in chronic kidney disease.

Production of Indole and Indole-Related Compounds by the Intestinal Microbiota and Consequences for the Host: The Good, the Bad, and the Ugly.

The intestinal microbiota metabolic activity towards the available substrates generates myriad bacterial metabolites that may accumulate in the luminal fluid. Among them, indole and indole-related compounds are produced by specific bacterial species from tryptophan. Although indole-related compounds are, first, involved in intestinal microbial community communication, these molecules are also active on the intestinal mucosa, exerting generally beneficial effects in different experimental situations. After absorption, indole is partly metabolized in the liver into the co-metabolite indoxyl sulfate. Although some anti-inflammatory actions of indole on liver cells have been shown, indoxyl sulfate is a well-known uremic toxin that aggravates chronic kidney disease, through deleterious effects on kidney cells. Indoxyl sulfate is also known to provoke endothelial dysfunction. Regarding the central nervous system, emerging research indicates that indole at excessive concentrations displays a negative impact on emotional behavior. The indole-derived co-metabolite isatin appears, in pre-clinical studies, to accumulate in the brain, modulating brain function either positively or negatively, depending on the doses used. Oxindole, a bacterial metabolite that enters the brain, has shown deleterious effects on the central nervous system in experimental studies. Lastly, recent studies performed with indoxyl sulfate report either beneficial or deleterious effects depending once again on the dose used, with missing information on the physiological concentrations that are reaching the central nervous system. Any intervention aiming at modulating indole and indole-related compound concentrations in the biological fluids should crucially take into account the dual effects of these compounds according to the host tissues considered.

Dip-and-Read, Organic Solvent-Compatible, Paper-Based Analytical Devices Equipped with Chromatographic Separation for Indole Analysis in Shrimp.

We developed an organic solvent-compatible paper-based analytical device (PAD) for the quantitative analysis of indole, which is an indicator of shrimp freshness. Although indole is insoluble in water, ethyl acetate is a suitable solvent to dissolve and extract indole from shrimp. The PADs are fabricated using a cutting method that allows the use of an organic solvent because no hydrophobic barrier is needed to form fluidic channels. Ehrlich's reagent consists of 4-(dimethylamino)benzaldehyde and p-dimethylaminobenzaldehyde and was deposited onto the reaction zone of the PAD followed by lamination to prevent evaporation of the ethyl acetate. Samples are introduced into the PAD via immersion in organic sample solutions. When the PAD is immersed into an indole solution of ethyl acetate in a closed bottle, the sample solution penetrates the channel of the PAD and successively flows into the detection zone to form a hydrophilic colored product. The PADs provide a linear relationship between the logarithm of the indole concentration and the color intensity within a range of 1.0-20 ppm with correlation coefficients of r2 > 0.99. The limits of detection and quantification are 0.36 and 0.71 ppm, respectively. Relative standard deviations for both the intraday (n = 2) and interday (n = 3) precision were less than 2.5%. In the indole analysis of shrimp, the PADs separated the interfering orange-colored astaxanthin in the extract from the colored product of indole via the paper chromatographic principle. We used the PADs to investigate the degradation of shrimp, and the results showed a rapid increase in the indole level after 7 days. High-performance liquid chromatography verified the accuracy of the PADs by showing good agreement with the obtained indole levels.

Detection of Bioactive Metabolites in Escherichia Coli Cultures Using Surface-Enhanced Raman Spectroscopy.

Detection of bioactive metabolites produced by bacteria is important for identifying biomarkers for infectious diseases. In this study, a surface-enhanced Raman spectroscopy (SERS)-based technique was developed for the detection of bioactive metabolite indole produced by Escherichia coli (E. coli) in biological media. The use of highly sensitive Au@Ag core-shell nanoparticles resulted in the detection of indole concentration as low as 0.0886 mM in standard solution. The supplementation of growth media with 5 mM of exogenous tryptophan resulted in the production of a maximum yield of indole of 3.139 mM by E. coli O157:H7 at 37 °C. The growth of bacterial cells was reduced from 47.73 × 108 to 1.033 × 106 CFU/mL when the cells were grown in 0 and 10 mM exogenous tryptophan, respectively. The amount of indole in the Luria–Bertani (LB) media had an inverse correlation with the growth of cells, which resulted in a three-log reduction in the colony-forming unit when the indole concentration in the media was 20 times higher than normal. This work demonstrates that SERS is an effective and highly sensitive method for rapid detection of bioactive metabolites in biological matrix.

Association between Smoking and Urine Indole Levels Measured by a Commercialized Test.

Indoles are formed from dietary tryptophan by tryptophanase-positive bacterium. A few amounts of indole are excreted in the urine. On the other hand, cigarette smoke contains indoles, which could also change the urine indole levels. This study sought to elucidate the relationship between urine indole levels and smoking habits. A total of 273 healthy men (46 ± 6 years old) were enrolled in the study. Fasting urine and blood samples were obtained in the morning. The indole concentration was measured by a commercialized kit with a modified Kovac’s reagent. The relationship with smoking status was evaluated. The median value of the urine indole test was 29.2 mg/L (interquartile range; 19.6–40.8). The urine indole level was significantly elevated in the smoking subjects (non-smoking group, 28.9 (20.9–39.1) mg/L, n = 94; past-smoking group, 24.5 (15.7–35.5) mg/L, n = 108; current-smoking group, 34.3 (26.9–45.0) mg/L, n = 71). In the current-smoking group, urine indole levels correlated with the number of cigarettes per day (ρ = 0.224, p = 0.060). A multivariate regression test with stepwise method revealed that the factors relating to urine indole level were current smoking (yes 1/no 0) (standardized coefficient β = 0.173, p = 0.004), blood urea nitrogen (β = 0.152, p = 0.011), and triglyceride (β = −0.116, p = 0.051). The result suggests that smoking is associated with increased urine indole levels. The practical test might be used as a screening tool to identify the harmful effect of smoking.

In vitro fermentation profiles of different soybean oligosaccharides and their effects on skatole production and cecal microbiota of broilers.

ObjectiveThe objective of this study was to investigate the in vitro fermentation profiles of different soybean oligosaccharides (SBOs) and their effects on skatole production and cecal microbiota of broilers.MethodsFive SBOs with varying main component contents were fermented using an in vitro batch incubation inoculated with broiler cecal microbiota. Gas production was recorded automatically, skatole, indole and short-chain fatty acids (SCFAs) were determined using high-performance liquid chromatography, and microbial changes were analyzed using 16S DNA gene sequencing.ResultsThe addition of SBOs increased (p<0.05) gas production, suggesting bacterial growth-stimulating activities. In addition, the concentrations of indole were significantly (p<0.05) decreased after SBO supplementation, and SBO III, with higher sucrose and stachyose contents, decreased (p<0.05) the skatole level. Our results also revealed that the fermentation of SBOs by cecal microbiota produced (p<0.05) SCFAs, which were dominated by propionic acid, butyrate acid and lactic acid compared to the control. In addition, SBO III increased (p<0.05) the abundance of Firmicutes and Subdoligranulum and decreased that of Bacteroides.ConclusionThese results suggest that SBOs with higher sucrose and stachyose contents are promising prebiotics in modulating gut microbiota and reducing odor emission in broilers.

Phenotypic causal networks between boar taint compounds measured in biopsies and carcasses

Context Boar taint compounds (androstenone, skatole and indole) can be measured in pig carcasses, after slaughter or, alternatively, in biopsies of subcutaneous adipose tissue from selected living pigs. Measuring these compounds via biopsy enables data collection in selected animals and high standardisation regarding tissue-collection procedures for phenotyping. Because different analytical methods can be used to measure boar taint compounds, it is important to better understand the relationship between boar taint compounds measured in biopsies and in carcasses. Aims This research aimed to identify the causal relationship and causal effects among boar taint compounds (androstenone, skatole and indole) measured in pig adipose tissue from carcasses and biopsies. Methods The concentrations of androstenone (AC), skatole (SC) and indole (IC) measured in adipose tissue from pig carcasses and the concentrations of androstenone (AB), skatole (SB) and indole (IB) measured in biopsies were used to fit a multi-trait Structural Equation Model (SEM) considering causal network graphs obtained via inductive causation algorithm with or without a priori information. Models were compared using the deviance information criterion (DIC). Key results The best DIC was obtained in a model with a causal structure built using a priori information; however, this model was considered inappropriate, because it returned several null genetic correlations among traits described as positively correlated. The best structure returned using only inductive causation algorithm was IB → SC ← AB ← AC ← SB: SC → IC, which was obtained with an 80–70% high-probability distribution interval. This model returned positive genetic correlations and improved goodness-of-fit compared with the multi-trait model in all cases. Several causal relationships among boar taint compounds in carcasses and biopsies were identified. Conclusion Boar taint compounds measured in biopsies have direct effects on boar taint compounds measured in carcasses. Implications Knowledge concerning the causal structure of boar taint compounds may be used in breeding programs, helping in the formulation of selection indexes and improving the ability for prediction and selection of this complex trait (boar taint).

Highly efficient extraction of indole from model wash oil by using environmentally benign deep eutectic solvents

Indole is an important organic synthesis intermediate derived from the wash oil fraction in coal tar. Deep eutectic solvents (DESs) have been widely used in the separation field due to their unique properties such as easy preparation, low cost, green and high extraction efficiency, and reusability. In this study, five DESs, including tetramethylammonium chloride/ethylene glycol (TMAC/EG), triethylmethylammonium chloride/ethylene glycol (TEMAC/EG), tetraethylammonium chloride/ ethylene glycol (TEAC/EG), tetrapropylammonium chloride/ethylene glycol (TPAC/EG), tetrabutylammonium chloride /ethylene glycol (TBAC/EG) were prepared and used to extract indole from model wash oil. The results showed that TPAC/EG exhibited the best extraction ability under the same conditions. Then, the main experimental parameters such as mass ratio of DES to model wash oil, extraction temperature, extraction time, standing time and initial concentration of indole, were investigated when using TPAC/EG as extractant. In optimal conditions (25℃, 5 min, mass ratio 1:8), the extraction efficiency, selectivity and distribution coefficient for indole were 90.51%, 79.21% and 61.12, respectively. Moreover, removing neutral oil with petroleum ether, the purity of indole further reached 96.63%, which was much higher than that in the previous studies reported. In addition, accessible and environmentally friendly water instead of diethyl ether was used to regenerate the DES. Reused four times, the TPAC/EG extraction ability for indole almost did not change. Finally, the mechanism of extraction process was investigated by FT-IR analysis. This work provides a new route for extraction and recovery indole from wash oil.

Nicotine Oral Administration Attenuates DSS-Induced Colitis Through Upregulation of Indole in the Distal Colon and Rectum in Mice.

Nicotine affects the gastrointestinal environment and modulates ulcerative colitis (UC). However, the associations among nicotine, gut metabolites, and UC are still largely unknown. We investigated whether orally administered nicotine affected gut metabolites and dextran sodium sulfate (DSS)-induced colitis. C57BL/6 male mice were orally administered nicotine solution in drinking water prior to inducing DSS-induced colitis. Short-chain fatty acids (SCFAs) and indole in gut contents and fecal samples were measured by GC-MS and hydroxylamine-based indole assays, respectively. Oral administration of nicotine increased indole concentration in feces, but, in contrast, SCFA values did not differ with nicotine administration. Indole levels were increased in the distal colon and rectum but not in the cecum and proximal colon. DSS-induced colitis was less severe clinically and histological changes were minimal in the rectum of orally nicotine-administered mice compared to mice drinking only water. 16S rRNA microbiome on the feces revealed an increasing in Clostridium and Porphyromonas in nicotine-administered mice. In conclusion, nicotine administration was associated with increased indole levels in the distal colon and rectum and attenuated DSS-induced colitis. Oral administration of nicotine may play a potential role in indole upregulation and prevention of UC.

Gut Microbiota and Their Derived Metabolites, a Search for Potential Targets to Limit Accumulation of Protein-Bound Uremic Toxins in Chronic Kidney Disease.

Chronic kidney disease (CKD) is characterized by gut dysbiosis with a decrease in short-chain fatty acid (SCFA)-producing bacteria. Levels of protein-bound uremic toxins (PBUTs) and post-translational modifications (PTMs) of albumin increase with CKD, both risk factors for cardiovascular morbidity and mortality. The relationship between fecal metabolites and plasma concentrations of PBUTs in different stages of CKD (n = 103) was explored. Estimated GFR tends to correlate with fecal butyric acid (BA) concentrations (rs = 0.212; p = 0.032), which, in its turn, correlates with the abundance of SCFA-producing bacteria. Specific SCFAs correlate with concentrations of PBUT precursors in feces. Fecal levels of p-cresol correlate with its derived plasma UTs (p-cresyl sulfate: rs = 0.342, p < 0.001; p-cresyl glucuronide: rs = 0.268, p = 0.006), whereas an association was found between fecal and plasma levels of indole acetic acid (rs = 0.306; p = 0.002). Finally, the albumin symmetry factor correlates positively with eGFR (rs = 0.274; p = 0.005). The decreased abundance of SCFA-producing gut bacteria in parallel with the fecal concentration of BA and indole could compromise the intestinal barrier function in CKD. It is currently not known if this contributes to increased plasma levels of PBUTs, potentially playing a role in the PTMs of albumin. Further evaluation of SCFA-producing bacteria and SCFAs as potential targets to restore both gut dysbiosis and uremia is needed.