iTRAQ-Based Quantitative Proteomic Analysis of Digestive Juice across the First 48 Hours of the Fifth Instar in Silkworm Larvae.

The silkworm is an oligophagous insect for which mulberry leaves are the sole diet. The nutrients needed for vital activities of the egg, pupal, and adult stages, and the proteins formed in the cocoon, are all derived from the larval stages. The silkworm feeds and grows quickly during the larval stages. In particular, the amount of leaf ingested and digested quickly increases from the ecdysis to the gluttonous stage in the fifth instar period. In this study, we used the iTRAQ proteomic technique to identify and analyze silkworm larval digestive juice proteins during this period. A total of 227 proteins were successfully identified. These were primarily serine protease activity, esterase activity, binding, and serine protease inhibitors, which were mainly involved in the digestion and overcoming the detrimental effects of mulberry leaves. Moreover, 30 genes of the identified proteins were expressed specifically in the midgut. Temporal proteomic analysis of digestive juice revealed developmental dynamic features related to molecular mechanisms of the principal functions of digesting, resisting pathogens, and overruling the inhibitory effects of mulberry leaves protease inhibitors (PIs) with a dynamic strategy, although overruling the inhibitory effects has not yet been confirmed by previous study. These findings will help address the potential functions of digestive juice in silkworm larvae.

Proteomics analysis of digestive juice from silkworm during Bombyx mori nucleopolyhedrovirus infection.

Previous studies have analyzed the midgut transcriptome and proteome after challenge with Bombyx mori nucleopolyhedrovirus (BmNPV), however little information is available on the digestive juice proteome after BmNPV challenge. This study investigated BmNPV infection-induced protein changes in the digestive juice of silkworms using shotgun proteomics and MS sequencing. From the digestive juice of normal third-day, fifth-instar silkworm larvae, 75 proteins were identified, 44 of which were unknown; from larvae 6 h after inoculation with BmNPV, 106 proteins were identified, of which 39 were unknown. After BmNPV challenge, more secreted proteins appeared that had antiviral and digestive features. GO annotation analysis clustered most proteins in the lumen into catalytic, binding, and metabolic processes. Numerous proteins were reported to have BmNPV interactions. Hsp70 protein cognate, lipase-1, and chlorophyllide A-binding protein precursor were upregulated significantly after BmNPV challenge. Levels of trypsin-like serine protease, beta-1,3-glucanase, catalase, and serine protease transcripts decreased or were not significantly change after BmNPV challenge. Taken together, these findings provided insights into the interaction between host and BmNPV and revealed potential functions of digestive juice after per os BmNPV infection.

Bioaccessibility of Glucosylceramide in Rice Based on the Cooking Condition and Cultivar.

Glucosylceramide (GlcCer), a major sphingolipid in plants, possesses various food functions, including improvement of intestinal impairments. This study evaluated rice cooking conditions and cultivars based on GlcCer levels transferred into the digestive juice using an in vitro digestion model to investigate the factors related to GlcCer availability. GlcCer levels transferred into the digestive juice were higher in rice gruel than in boiled rice. The GlcCer levels in the digestive juice of boiled rice varied based on the rice cultivar, whereas those in rice gruel had no difference. Thus, GlcCer in rice was not fully utilized via digestion. Further, bioaccessibility was related to the amylose ratio and added water content.

Insights into midgut cell types and their crucial role in antiviral immunity in the lepidopteran model Bombyx mori.

The midgut, a vital component of the digestive system in arthropods, serves as an interface between ingested food and the insect's physiology, playing a pivotal role in nutrient absorption and immune defense mechanisms. Distinct cell types, including columnar, enteroendocrine, goblet and regenerative cells, comprise the midgut in insects and contribute to its robust immune response. Enterocytes/columnar cells, the primary absorptive cells, facilitate the immune response through enzyme secretions, while regenerative cells play a crucial role in maintaining midgut integrity by continuously replenishing damaged cells and maintaining the continuity of the immune defense. The peritrophic membrane is vital to the insect's innate immunity, shielding the midgut from pathogens and abrasive food particles. Midgut juice, a mixture of digestive enzymes and antimicrobial factors, further contributes to the insect's immune defense, helping the insect to combat invading pathogens and regulate the midgut microbial community. The cutting-edge single-cell transcriptomics also unveiled previously unrecognized subpopulations within the insect midgut cells and elucidated the striking similarities between the gastrointestinal tracts of insects and higher mammals. Understanding the intricate interplay between midgut cell types provides valuable insights into insect immunity. This review provides a solid foundation for unraveling the complex roles of the midgut, not only in digestion but also in immunity. Moreover, this review will discuss the novel immune strategies led by the midgut employed by insects to combat invading pathogens, ultimately contributing to the broader understanding of insect physiology and defense mechanisms.

The influence of gastrointestinal pH on speciation of copper in simulated digestive juice.

Speciation can provide knowledge about absorption, reactivity to binding sites, bioavailability, toxicity, and excretion of elements. In this study, the speciation of copper in different model solutions under the influence of gastrointestinal (GI) pH was studied by ion selective electrode (ISE) and inductively coupled plasma optical emission spectrometry (ICP OES). It was found that the electrode response (mV) against Cu2+ decreased with the increase in pH and dropped to the lowest point at pH 7.5 in all model solutions. When amino acids and organic acids were present, the ratio of filtered copper (0.45 µm, pH 7.5) was more than 90%. When casein was present, whey protein, pancreatin, and starch were added, and the ratio of filtered copper was 85.6 ± 0.3, 56.7 ± 8.8, 38.5 ± 5.1, and 1.0 ± 0.3%, respectively. When there is not enough organic ligand, excessive copper will form copper hydroxide precipitation with the increase in pH, but it got the highest electrode response (mV) against Cu2+. From this study, it can be concluded that the speciation of copper in GI tract is strongly influenced by the pH and the composition of food. When there are few ligands coexisting in the GI tract, the concentration of copper ion may be relatively high.

Potential Hypolipidemic Effects of Banana Condensed Tannins Through the Interaction with Digestive Juice Components Related to Lipid Digestion.

An in vitro intestinal model was used to evaluate the impact of banana condensed tannins (BCT) on the digestion of lipids (fat and cholesterol). BCT significantly suppressed the digestion of fat and cholesterol by interacting with digestive juice components. The interactions of BCT with a digestive juice mixture and its components (including bile acid, lipase, cholesterol esterase, CaCl2, NaCl, and cholesterol) were analyzed using turbidity, isothermal titration calorimetry, particle size distribution, zeta potential, and molecular docking analyses. The results showed that BCT reduced the digestion of lipids mainly via interaction with lipase, cholesterol esterase, bile acid, and cholesterol. Electrostatic CT-calcium ion complexes might reduce the extent of lipid digestion by decreasing the surface area of the lipid droplets exposed to the enzymes. This research provides valuable insights into the molecular mechanisms of the interaction of BCT with digestive juice components related to lipid digestion that may affect the rate and extent of lipid digestion.

Quantitative label-free proteomic analysis reveals differentially expressed proteins in the digestive juice of resistant versus susceptible silkworm strains and their predicted impacts on BmNPV infection.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a major pathogen causing severe economic loss. Previous studies have revealed that some proteins in silkworm digestive juice show antiviral activity. In this study, antiviral activity examination of different resistant strains showed that the digestive juice of the resistant strain (A35) had higher inhibition to virus than the susceptible strain (P50). Subsequently, the label-free quantitative proteomics was used to study the midgut digestive juice response to BmNPV infection in P50 and A35 strains. A total of 98 proteins were identified, of which 80 were differentially expressed proteins (DEPs) with 54 enzymes and 26 nonenzymatic proteins by comparing the proteomes of infected and non-infected P50 and A35 silkworms. These DEPs are mainly involved in metabolism, proteolysis, neuroactive ligand receptor interaction, starch and sucrose metabolism and glutathione metabolism. After removing the genetic background and individual immune stress response proteins, 9 DEPs were identified potentially involved in resistance to BmNPV. Further studies showed that a serine protease, an alkaline phosphatase and serine protease inhibitor 2 isoform X1 were differentially expressed in A35 compared to P50 or post BmNPV infection. Taken together, these results provide insights into the potential mechanisms for silkworm digestive juice to provide resistance to BmNPV infection. Signifcance: Bombyx mori nucleopolyhedrovirus (BmNPV) is highly pathogenic, which has a great impact on the sericulture. BmNPV entered the midgut lumen and exposed to digestive juices after oral infection. Previous studies have revealed that some proteins in silkworm digestive juice show antiviral activity, however, current information on the digestive juice proteome of high resistant silkworm strain after BmNPV challenge compared to susceptible strain is incomprehensive. Here, we combined label-free quantification method, bioinformatics, RT-qPCR and western blot analysis and found that BmNPV infection causes some protein changes in the silkworm midgut digestive juice. The DEPs were identified in the digestive juices of different resistant strains following BmNPV infection, and screened out some proteins potentially related to resistance to BmNPV. Three important differentially expression proteins were validated by independent approaches. These findings uncover the potential role of silkworm digestive juice in providing resistance to BmNPV and supplemented the profile of the proteome of the digestive juices in B. mori.

Effect of Simultaneous Snail Slime-aided Degradation and Yeast Fermentation on Terpenoid Composition of Plantain Pseudostem Waste.

BACKGROUND: In this study, local sustainable enzyme sources involving excised digestive juice of African land snail and yeast were utilized to achieve the simultaneous saccharification (SSF) and fermentation (SSF) of plantain pseudostem (PPS) waste, and afterwards their effects on terpenoids using gas chromatography coupled to a flame ionization detector (GC-FID), were examined. METHODS: The most abundant terpenoids were found in the order α-pinene > borneol > camphor > humulene > ß-caryophellene, while the least in abundance were cis ocimene (8.78x10-6 mg/100g), and cyperene (1.81x10-5 mg/100g). The application of exclusive fermentation and SSF respectively elevated azuluene by 95.46 and 99.6%, while pinene-2-ol was elevated by 83.02 and 98.57%, respectively. RESULTS: Both exclusive fermentation and SSF had no effect on myrcene, cyperene, ethyl cinnamate, germacrene b, valencene, beta selinene, aromadendrene, and taraxerol, while the degree of degradation of some of the terpenoids by both processes was respectively as follows; gama muurolene (100%), ß-caryophyllene (97.60 and 93.14%), α-terpinenyl acetate (91.95 and 83.16%), geranyl acetate (94.81 and 43.87%), and terpinen-4-ol (94.40 and 57.00%). CONCLUSION: The findings of this study encourage the imminent application of simultaneous saccharification and fermentation for the enhancement of bioactivities of terpenoids.

Probiotic Properties of Lactobacillus Plantarum LRCC5193, a Plant-Origin Lactic Acid Bacterium Isolated from Kimchi and Its Use in Chocolates.

This study involves an investigation of the probiotic properties of lactic acid bacteria isolated from Kimchi, and their potential applications in chocolate. Lactobacillus plantarum-LRCC5193 (LP-LRCC5193) demonstrated a significantly higher degree of heat, acid, and bile acid tolerance compared to other Kimchi isolates. The intestinal adhesion assay also revealed that 84.2 log percentage of LP-LRCC5193 adhered to the Caco-2 cells after 2 h of incubation. Furthermore, the lyophilized LP-LRCC5193 maintained 92.9 log percentage and 97.2 log percentage survival rate within artificial stomach juice (pH 2.5, pepsin 0.04%) and artificial intestinal juice (oxgall 0.5%, trypsin 0.04%, and pancreatin 0.04%), respectively. Meanwhile, we also found that lyophilized LP-LRCC5193 incorporated in chocolate exhibited significantly higher survivability than lyophilized LP-LRCC5193 in both artificial gastric and intestinal juice under 1 to 3 hr incubation, where the survivability was within the range of 96.3 to 98.5 log percentage, and 98.8 to 98.9 log percentage, respectively. A 6-month storage test further revealed that LP-LRCC5193 demonstrated higher stability than the lyophilized LP-LRCC5103 in 3 different temperature ranges, where the final survival rates were 97.2 log percentage (20 °C), 89.2 log percentage (33 °C), and 94.4 log percentage (15 to 30 °C/wk). Altogether, our data suggest that chocolate can be used as a tasty delivery vehicle for delivering putative probiotic strain, LP-LRCC5193 to the gastrointestinal tract. PRACTICAL APPLICATION: Lactobacillus plantarum LRCC5193 (LP-LRCC5193) isolated from Kimchi demonstrated high stability under gastrointestinal environmental stresses and good adhesion to the intestinal epithelial cells in vitro. In addition, LP-LRCC5193 containing chocolates remained highly stable after storage at room temperature for 6 months. Chocolate containing LP-LRCC5193 can thus be considered a promising probiotic delivery system.

Bioaccessibility of phenolic compounds, lutein, and bioelements of preparations containing Chlorella vulgaris in artificial digestive juices.

Chlorella vulgaris Beijerinck is a spherical, green alga belonging to the genus Chlorella and family Chlorellaceae. It has high nutritional value and shows multiple biological effects. Dietary supplements that contain extracts of C. vulgaris are sold in the form of tablets, capsules, powders, and aqueous solutions. To the best of our knowledge, this is the first study to determine the content of bioelements (zinc, iron, and magnesium), phenolic compounds, and lutein before and after incubation with artificial digestive juices from preparations containing C. vulgaris. In this study, we used commercial preparations in the form of powder and tablets. The samples were incubated in artificial gastric juice and then in artificial intestinal juice for 30 and 90 min. The contents of bioelements were determined by using the flame atomic absorption spectrometric method. Lutein and phenolic compounds were analyzed by high-pressure liquid chromatography. We also aimed to evaluate the quality of chlorella-containing formulations by using the methods described in the European Pharmacopoeia 8th edition. According to the results, the preparations containing C. vulgaris demonstrated the presence of phenolic compounds and lutein. Therefore, daily supplementation of preparations containing C. vulgaris substantiates its usefulness for humans. The qualitative composition of the examined organic substances and bioelements was found to be in accordance with the manufacturer's declarations on the packaging containing C. vulgaris compared with the control samples; however, the contents of bioelements were found to be negligible after incubation with artificial digestive juices. This shows that the examined preparations containing C. vulgaris are not good sources of bioelements such as zinc, iron, or magnesium.

Human Gastrointestinal Metabolism of the Cistanches Herba Water Extract in Vitro: Elucidation of the Metabolic Profile Based on Comprehensive Metabolite Identification in Gastric Juice, Intestinal Juice, Human Intestinal Bacteria, and Intestinal Microsomes.

Cistanches Herba is taken orally as a health food supplement and medicinal plant in Asian countries. It consists of the stems of Cistanche deserticola (CD) and Cistanche tubulosa (CT). The gastrointestinal metabolism of the multiple components contained in Cistanches Herba is crucial for the discovery of bioactive constituents. This study aims to elucidate the comprehensive metabolic profile of the Cistanches Herba water extract by simulating human gastrointestinal metabolism in vitro independently and sequentially using four models: gastric juice, intestinal juice, human intestinal bacteria, and human intestinal microsomes. A total of 35 and 18 metabolites were characterized from CD and CT water extracts, respectively. These metabolites were formed through reduction, methylation, dimethylation, deglycosylation, decaffeoyl, derhamnose, dehydrogenation, and glucuronidation. The difference in metabolites of the Cistanches Herba water extract and single compounds and the difference in metabolites of CD and CT water extracts were caused by the oligosaccharides and polysaccharides in Cistanches Herba.

In vitro culture of Boletus badius as a source of indole compounds and zinc released in artificial digestive juices.

The objective of this study was to obtain the in vitro cultures of Boletus badius under controlled conditions and investigate the release of indole compounds and zinc from the mycelium of B. badius to artificial digestive juices under conditions similar to those in the human gastrointestinal tract. Biomass was obtained from cultures grown using both only the Oddoux medium as well as the same medium with added zinc hydroaspartate and zinc sulfate. The release of 5-hydroxy-L-tryptophan, L-tryptophan, and serotonin from the B. badius biomass extracts to the artificial digestive juices was determined. Differential pulse anodic stripping voltammetry was used to demonstrate that zinc is released from each of the extracted materials. The total amount of zinc in the materials under study was estimated to be between 7.12 and 44.15 mg/100 g dry weight. It was demonstrated that in vitro cultures of B. badius grown using appropriately selected media may supplement zinc and indole compounds.

Effects of phoxim on nutrient metabolism and insulin signaling pathway in silkworm midgut.

Silkworm (Bombyx mori) is an important economic insect. Each year, poisoning caused by phoxim pesticide leads to huge economic losses in sericulture in China. Silkworm midgut is the major organ for food digestion and nutrient absorption. In this study, we found that the activity and expression of nutrition metabolism-related enzymes were dysregulated in midgut by phoxim exposure. DGE analysis revealed that 40 nutrition metabolism-related genes were differentially expressed. qRT-PCR results indicated that the expression levels of insulin/insulin growth factor signaling (IIS) pathway genes Akt, PI3K, PI3K60, PI3K110, IRS and PDK were reduced, whereas PTEN's expression was significantly increased in the midgut at 24 h after phoxim treatment. However, the transcription levels of Akt, PI3K60, PI3K110, IRS, InR and PDK were elevated and reached the peaks at 48 h, which were 1.48-, 1.35-, 1.21-, 2.24-, 2.89-, and 1.44-fold of those of the control, respectively. At 72 h, the transcription of these genes was reduced. Akt phosphorylation level was increasing along with the growth of silkworms in the control group. However, phoxim treatment led to increased Akt phosphorylation that surged at 24 h but gradually decreased at 48 h and 72 h. The results indicated that phoxim dysregulated the expression of IIS pathway genes and induced abnormal nutrient metabolism in silkworm midgut, which may be the reason of the slow growth of silkworms.

The closed re-infusion of intestinal juice for intestinal fistula / 肠外与肠内营养

Objective: To investigate the influence of the closed re-infusion of intestinal juice and EN on the treatment of intestinal fistula.Methods: The study group was composed of 21 patients of intestinal fistula from October 2004 to December 2006.The fistulae was in the 50 cm of the proximate jejunum and caused by trauma and operatin.The leak amount of intestinal juice was more than 1 000 mL.The protein level in serum and the absorption rates of the glucose,fat and nitrogen after the reinfusion of the intestinal juice were measured.Resutls: The protein level in serum and the absorption rates of the glusose,fat and nitrogen was increased after the re-infusion of the intestinal juice.Conclusion: The absorption of the glucose,fat and nitrogen in EN and the protein level in serum can be increased after the closed re-infusion of the intestinal juice.

Differentiation of CEA and CEA-Related Substances by Western Blot Technique in Biliary and Gastric Diseases / 中国普外基础与临床杂志

Objective To eliminate the interference of CEA-related substances in CEA measurement and increase the specificity of CEA in the detection of malignant digestive diseases.Methods CEA level of peripheral blood and digestive juice(bile,gastric juice) from patients with benign or malignant digestive diseases was measured by ELISA,and semi-dry electrophoretic transfer method of Western blot technique to distinguish CEA and CEA-related substances.Results In malignant diseases,the CEA level of digestive juice was significantly higher than that in the blood,and there was no difference of CEA level in digestive juice and blood in benign diseases.Meanwhile,the CEA level of digestive juice and blood in malignant diseases were significantly higher than that in benign diseases.A specific band(molecular weight about 210?10~3) was detected in all malignant diseases except four cases whose CEA level was too low(less than 5 ?g/L),whereas no one of benign diseases had this specific band no matter how high or low the CEA level was.Conclusion The specificity of CEA detection in malignant digestive diseases can be improved by using digestive juice as sample and combining with Western blot technique.