Caco-2 Monolayer as a Model of the Intestinal Barrier: Permeability of Magnesium Salts.

Magnesium, an essential mineral for various physiological functions, is subject to tight regulation within the body. Understanding its absorption across epithelial cell monolayers is crucial for optimizing dietary magnesium intake and therapeutic strategies. The Caco-2 monolayer model, widely recognized for its relevance to the human intestinal epithelium, provides a suitable platform for this investigation. This protocol covers the step-by-step procedures for the cultivation of Caco-2 monolayer preparation of transwell systems. It provides guidance on the setup of magnesium transport experiments, which involve the application of magnesium salts to the apical side of the Caco-2 monolayer and monitoring their transport to the basolateral side.

Evaluating intestinal absorption of peptide Met-Lys-Pro in casein hydrolysate using Caco-2 and human iPS cell-derived small intestinal epithelial cells.

High blood pressure is a major risk factor for cardiovascular disease. Our previous study confirmed that daily intake of casein hydrolysate that contained Met-Lys-Pro (MKP) can safely lower mildly elevated blood pressure. The present study aimed to evaluate the intestinal absorption differences between peptide MKP as a casein hydrolysate and synthetic MKP alone using Caco-2 cells and human iPS cell-derived small intestinal epithelial cells (hiSIECs). MKP was transported intact through Caco-2 cells and hiSIECs with permeability coefficient (Papp) values of 0.57 ± 0.14 × 10-7 and 1.03 ± 0.44 × 10-7 cm/s, respectively. This difference in Papp suggests differences in the tight junction strength and peptidase activity of each cell. Moreover, the transepithelial transport and residual ratio of intact MKP after adding casein hydrolysate containing MKP was significantly higher than that after adding synthetic MKP alone, suggesting that other peptides in casein hydrolysate suppressed MKP degradation and increased its　transport. These findings suggest that hiSIECs could be useful for predicting the human intestinal absorption of bioactive peptides; ingesting MKP as a casein hydrolysate may also improve MKP bioavailability.

Comparative Evaluation of Micellization and Cellular Uptake of ß-Carotene Affected by Flavonoids.

Based on in vitro digestion, micellar synthesis, and Caco-2 cell model, this study investigated the effects of typical flavonoids in citrus (naringenin, naringin, hesperetin, hesperidin, quercetin, and rutin) at different doses on the micellization and cellular uptake of ß-carotene. In in vitro digestion, low-dose flavonoids enhanced ß-carotene bioaccesssibility by regulating the stability and dispersibility of the intestinal medium, particularly quercetin, which significantly increased the bioaccessibility by 44.6% (p < 0.05). Furthermore, naringenin, hesperetin, hesperidin, and quercetin enhanced the micellar incorporation rate of ß-carotene; however, naringin and rutin exhibited an opposite effect, particularly naringin, which significantly reduced it by 71.3% (p < 0.05). This phenomenon could be attributed to the high solubility of naringin and rutin in micelles, resulting in a competitive inhibitory effect on ß-carotene. Besides, all treatments significantly enhanced ß-carotene cellular uptake (p < 0.05) by promoting the expression of scavenger receptor class B type I and Niemann-Pick C1-Like 1.

Bioaccessibility of iron in developed pectin iron complex using Citrus limon Burm F. peels subjected to in-vitro gastro-pancreatic digestion.

Pectin from the citrus peel waste has novel applications in food and biomedical industries. The present work focused on addressing iron deficiency, which is a global health concern, by developing a functional ingredient using pectin extracted from Assam lemon (Citrus limon Burm. F) and supplementing iron via the pectin­iron complex (PIC). Extracted pectin was incubated with iron chloride hexahydrate (0.90-1.80 mM) for 180 h to optimize the complexation conditions, with the optimal concentration being 1.36 mM. The iron bioavailability and its absorption in the PIC was assessed using in-vitro simulation digestion and Caco-2 cell monolayers. The bioaccessible form of iron in the developed PIC during the intestinal phase was 5.34 ± 0.16%, which was negligible in pectin. The absorption of bioaccessible iron in the PIC was found to be 2.93 ± 0.03%. The results demonstrated that PIC could reduce iron deficiency and increase fibre intake, leading to several health benefits.

Short Caco-2 model for evaluation of drug permeability: A sodium valerate-assisted approach.

The human colorectal adenocarcinoma cell line Caco-2, widely used for studying intestinal drug permeability, is typically grown on permeable filter supports and matures in 21 days with frequent media changes. The process is labor-intensive, prone to contamination, and has low throughput, contributing to the overall high utilization cost. Efforts to establish a low-cost, high-throughput, and short-duration model have encountered obstacles, such as weaker tight junctions causing monolayer leaks, incomplete differentiation resulting in low transporter expression, intricate and challenging protocols, and cytotoxicity, limiting the usability. Hence, this study aimed to develop a low-cost, efficient, and short-duration model by addressing the aforementioned concerns by customizing the media and finding a safe differentiation inducer. We generated a new rapid model using sodium valerate, which demonstrated sufficient transporter activity, improved monolayer integrity, and higher levels of differentiation markers than the 21-day model. Furthermore, this model exhibited consistent and reliable results when used to evaluate drug permeability over multiple days of repeated use. This study demonstrates the potential of a sodium valerate-assisted abbreviated model for drug permeability assessment with economic and practical advantages.

Emamectin benzoate-induced toxicity affects intestinal epithelial integrity involving apoptosis.

The frequency presence of emamectin benzoate in agricultural production highlights the need for studying their toxicity against human intestinal epithelial barrier (IEB). Herein, we combined a Caco-2 cell model with transcriptome analysis to assess the intestinal toxicity of emamectin benzoate and its disease-causing potential. Results showed that the half maximal inhibitory concentration (IC50) of emamectin benzoate on Caco-2 cell viability after 24, 48, and 72 h of exposure were 18.1, 9.9, and 8.3 µM, respectively. Emamectin benzoate exposure enhanced the Caco-2 monolayer paracellular permeability, damaged the IEB, and increased cellular apoptosis. Key driver gene analysis of 42 apoptosis - related DEGs, identified 10 genes (XIAP, KRAS, MCL1, NRAS, PIK3CA, CYCS, MAPK8, CASP3, FADD, and TNFRSF10B) with the strongest correlation with emamectin benzoate - induced apoptosis. Transcriptomics identified 326 differentially expressed genes (DEGs, 204 upregulated and 122 downregulated). The functional terms of neurodegeneration - multiple diseases was enriched with the most number of DEGs, and the Parkinson disease pathway had the highest enrichment degree. Our findings provided support for environmental toxicology studies and the health risk assessment of emamectin benzoate.

A Machine Learning Approach for Predicting Caco-2 Cell Permeability in Natural Products from the Biodiversity in Peru.

BACKGROUND: Peru is one of the most biodiverse countries in the world, which is reflected in its wealth of knowledge about medicinal plants. However, there is a lack of information regarding intestinal absorption and the permeability of natural products. The human colon adenocarcinoma cell line (Caco-2) is an in vitro assay used to measure apparent permeability. This study aims to develop a quantitative structure-property relationship (QSPR) model using machine learning algorithms to predict the apparent permeability of the Caco-2 cell in natural products from Peru. METHODS: A dataset of 1817 compounds, including experimental log Papp values and molecular descriptors, was utilized. Six QSPR models were constructed: a multiple linear regression (MLR) model, a partial least squares regression (PLS) model, a support vector machine regression (SVM) model, a random forest (RF) model, a gradient boosting machine (GBM) model, and an SVM-RF-GBM model. RESULTS: An evaluation of the testing set revealed that the MLR and PLS models exhibited an RMSE = 0.47 and R2 = 0.63. In contrast, the SVM, RF, and GBM models showcased an RMSE = 0.39-0.40 and R2 = 0.73-0.74. Notably, the SVM-RF-GBM model demonstrated superior performance, with an RMSE = 0.38 and R2 = 0.76. The model predicted log Papp values for 502 natural products falling within the applicability domain, with 68.9% (n = 346) showing high permeability, suggesting the potential for intestinal absorption. Additionally, we categorized the natural products into six metabolic pathways and assessed their drug-likeness. CONCLUSIONS: Our results provide insights into the potential intestinal absorption of natural products in Peru, thus facilitating drug development and pharmaceutical discovery efforts.

Qualification of In Vitro Dissolution Absorption System 2 (IDAS2) with Caco-2 and MDCK Cell Monolayers: Dose Sensitivity Study Using BCS Class I and III Drugs.

This study aimed to validate the In vitro Dissolution Absorption System 2 (IDAS2) containing a biological barrier of Caco-2 or Madin-Darby canine kidney (MDCK) cell monolayer through dose sensitivity studies. Metoprolol and propranolol were selected as Biopharmaceutics Classification System (BCS) Class I model drugs, and atenolol as a Class III model drug. The IDAS2 is comprised of a dissolution vessel (500 mL) and two permeation chambers (2 × 8.0 mL) mounted with Caco-2 or MDCK cell monolayer. One or two immediate-release tablet(s) of the model drug were added to the dissolution vessel, and the time profiles of dissolution and permeation were observed. Greater than 85% of metoprolol and propranolol (tested at two dosing concentrations) were dissolved by 15 min, and all drugs were fully dissolved by 30 min. All three drugs were more permeable across Caco-2 cells than MDCK cells with a linear increase in permeation across both cells at both dose concentrations. Thus, the dose sensitivity of the IDAS2 was demonstrated using both cell barriers. These results indicate a successful qualification of IDAS2 for the development/optimization of oral formulations and that MDCK cells can be utilized as a surrogate for Caco-2 cells.

Whey Protein Sodium-Caseinate as a Deliverable Vector for EGCG: In Vitro Optimization of Its Bioaccessibility, Bioavailability, and Bioactivity Mode of Actions.

Epigallocatechin gallate (EGCG), the principal catechin in green tea, exhibits diverse therapeutic properties. However, its clinical efficacy is hindered by poor stability and low bioavailability. This study investigated solid particle-in-oil-in-water (S/O/W) emulsions stabilized by whey protein isolate (WPI) and sodium caseinate (NaCas) as carriers to enhance the bioavailability and intestinal absorption of EGCG. Molecular docking revealed binding interactions between EGCG and these macromolecules. The WPI- and NaCas-stabilized emulsions exhibited high encapsulation efficiencies (>80%) and significantly enhanced the bioaccessibility of EGCG by 64% compared to free EGCG after simulated gastrointestinal digestion. Notably, the NaCas emulsion facilitated higher intestinal permeability of EGCG across Caco-2 monolayers, attributed to the strong intermolecular interactions between caseins and EGCG. Furthermore, the emulsions protected Caco-2 cells against oxidative stress by suppressing intracellular reactive oxygen species generation. These findings demonstrate the potential of WPI- and NaCas-stabilized emulsions as effective delivery systems to improve the bioavailability, stability, and bioactivity of polyphenols like EGCG, enabling their applications in functional foods and nutraceuticals.

Mitigating cadmium exposure risk in rice with foliar nano-selenium: Investigations through Caco-2 human cell line in-vivo bioavailability assay.

The contamination of paddy fields by cadmium and lead is a major issue in China. The consumption of rice grown in heavy metals contaminated areas poses severe health risks to humans, where bioavailability and bioaccessibility remains the critical factor for risk determination. Selenium nanoparticles (Se-NPs) can mitigate the toxicity of heavy metals in plants. However, there exists limited information regarding the role of Se-NPs in dictating cadmium (Cd) toxicity in rice for human consumption. Moreover, the impact of Se-NPs under simultaneous field and laboratory controlled conditions is rarely documented. To address this knowledge gap, a field experiment was conducted followed by laboratory scale bioavailability assays. Foliar application of Se-NPs and selenite (at 5, 10 mg L-1) was performed to assess their efficiency in lowering Cd accumulation, promoting Se biofortification in rice grains, and evaluating Cd exposure risk from contaminated rice. Obtained results indicate that foliar treatments significantly reduced the heavy metal accumulation in rice grains. Specifically, Se-NP 10 mg L-1 demonstrated higher efficiency, reducing Cd and Pb by 56 and 32 % respectively. However, inconsistent trends for bioavailable Cd (0.03 mg kg-1) and bioaccessible (0.04 mg kg-1) were observed while simulated human rice intake. Furthermore, the foliage application of Se-NPs and selenite improved rice quality by elevating Se, Zn, Fe, and protein levels, while lowering phytic acid content in rice grains. In summary, this study suggests the promising potential of foliage spraying of Se-NPs in lowering the health risks associated with consuming Cd-contaminated rice.

Single-cell calcium monitoring of Caco-2 cell co-cultured with intestinal microbiome through carbon fiber based potentiometric microelectrode.

The Caco-2 cells were used as intestinal epithelial cell model to illustrate the hyperuricemia (HUA) mechanism under the co-culture of the imbalanced intestinal microbiome in this work. The uric acid (UA) concentration in the HUA process was monitored, and could be up to 425 µmol/L at 8 h co-cultured with the imbalanced intestinal microbiome. Single-cell potentiometry based on ion-selective microelectrode was used to study extracellular calcium change, which is hypothesized to play an important role in the UA excretion. The potential signal of the calcium in the extremely limited microenvironment around single Caco-2 cell was recorded through the single-cell analysis platform. The potential signal of sharp decrease and slow increase followed within a few seconds indicates the sudden uptake and gradually excretion process of calcium through the cell membrane. Moreover, the value of the potential decrease increases with the increase of the time co-cultured with the imbalanced intestinal microbiome ranging from 0 to 8 h. The Ca2+ concentration around the cell membrane could decrease from 1.3 mM to 0.4 mM according to the potential decrease of 27.0 mV at the co-culture time of 8 h. The apoptosis ratio of the Caco-2 cells also exhibits time dependent with the co-culture of the imbalanced intestinal microbiome, and was 39.1 ± 3.6 % at the co-culture time of 8 h, which is much higher than the Caco-2 cells without any treatment (3.9 ± 2.9 %). These results firstly provide the links between the UA excretion with the apoptosis of the intestinal epithelial cell under the interaction of the imbalanced intestinal microbiome. Moreover, the apoptosis could be triggered by the calcium signaling.

Absorption and intracellular accumulation of food-borne dicarbonyl precursors of advanced glycation end-product in a Caco-2 human cell transwell model.

This study aimed to better understand whether and how the reactive 1,2-dicarbonyl precursors of advanced glycation end products (AGEs), glyoxal (GO) and methylglyoxal (MGO), cross the intestinal barrier by studying their transport in the in vitro Caco-2 transwell system. The results reveal that GO, MGO and Nε-(carboxymethyl)lysine (CML), the latter studied for comparison, are transported across the intestinal cell layer via both active and passive transport and accumulate in the cells, albeit all to a limited extent. Besides, the transport of the dicarbonyl compounds was only partially affected by the presence of amino acids and protein, suggesting that scavenging by a food matrix will not fully prevent their intestinal absorption. Our study provides new insights into the absorption of the two major food-borne dicarbonyl AGE precursors and provides evidence of their potential systemic bioavailability but also of factors limiting their contribution to the overall exposome.

Metabolomic insights into the profile, bioaccessibility, and transepithelial transport of polyphenols from germinated quinoa during in vitro gastrointestinal digestion/Caco-2 cell transport, and their prebiotic effects during colonic fermentation.

The health-promoting activities of polyphenols and their metabolites originating from germinated quinoa (GQ) are closely related to their digestive behavior, absorption, and colonic fermentation; however, limited knowledge regarding these properties hinder further development. The aim of this study was to provide metabolomic insights into the profile, bioaccessibility, and transepithelial transport of polyphenols from germinated quinoa during in vitro gastrointestinal digestion and Caco-2 cell transport, whilst also investigating the changes in the major polyphenol metabolites and the effects of prebiotics during colonic fermentation. It was found that germination treatment increased the polyphenol content of quinoa by 21.91%. Compared with RQ group, 23 phenolic differential metabolites were upregulated and 47 phenolic differential metabolites were downregulated in GQ group. Compared with RQ group after simulated digestion, 7 kinds of phenolic differential metabolites were upregulated and 17 kinds of phenolic differential metabolites were downregulated in GQ group. Compared with RQ group after cell transport, 7 kinds of phenolic differential metabolites were upregulated and 9 kinds of phenolic differential metabolites were downregulated in GQ group. In addition, GQ improved the bioaccessibilities and transport rates of various polyphenol metabolites. During colonic fermentation, GQ group can also increase the content of SCFAs, reduce pH value, and adjust gut microbial populations by increasing the abundance of Actinobacteria, Bacteroidetes, Verrucomicrobiota, and Spirochaeota at the phylum level, as well as Bifidobacterium, Megamonas, Bifidobacterium, Brevundimonas, and Bacteroides at the genus level. Furthermore, the GQ have significantly inhibited the activity of α-amylase and α-glucosidase. Based on these results, it was possible to elucidate the underlying mechanisms of polyphenol metabolism in GQ and highlight its beneficial effects on the gut microbiota.

Antioxidant and anti-inflammatory function of Eupatorium adenophora Spreng leaves (EASL) on human intestinal Caco-2 cells treated with tert-butyl hydroperoxide.

Chronic non-communicable diseases (CNCDs) pose a significant public health challenge. Addressing this issue, there has been a notable breakthrough in the prevention and mitigation of NCDs through the use of antioxidants and anti-inflammatory agents. In this study, we aim to explore the effectiveness of Eupatorium adenophora Spreng leaves (EASL) as an antioxidant and anti-inflammatory agent, and its potential applications. To construct a cellular model of oxidative damage and inflammation, Caco-2 cells were treated with tert-butyl hydroperoxide (t-BHP). The biocompatibility of EASL-AE with Caco-2 cells was assessed using the MTT assay, while compatibility was further verified by measuring LDH release and the protective effect against oxidative damage was also assessed using the MTT assay. Additionally, we measured intracellular oxidative stress indicators such as ROS and 8-OHdG, as well as inflammatory pathway signalling protein NFκB and inflammatory factors TNF-α and IL-1ß using ELISA, to evaluate the antioxidant and anti-inflammatory capacity of EASL-AE. The scavenging capacity of EASL-AE against free radicals was determined through the DPPH Assay and ABTS Assay. Furthermore, we measured the total phenolic, total flavonoid, and total polysaccharide contents using common chemical methods. The chemical composition of EASL-AE was analyzed using the LC-MS/MS technique. Our findings demonstrate that EASL-AE is biocompatible with Caco-2 cells and non-toxic at experimental levels. Moreover, EASL-AE exhibits a significant protective effect on Caco-2 cells subjected to oxidative damage. The antioxidant effect of EASL-AE involves the scavenging of intracellular ROS, while its anti-inflammatory effect is achieved by down-regulation of the NFκB pathway. Which in turn reduces the release of inflammatory factors TNF-α and IL-1ß. Through LC-MS/MS analysis, we identified 222 compounds in EASL-AE, among which gentianic acid, procaine and L-tyrosine were the compounds with high antioxidant capacity and may be the effective constituent for EASL-AE with antioxidant activity. These results suggest that EASL-AE is a natural and high-quality antioxidant and anti-inflammatory biomaterial that warrants further investigation. It holds great potential for applications in healthcare and other related fields.

Use of human Caco-2 cells and HPAE-PAD for α-glucosidase assay.

To measure α-glucosidase activity, rat intestinal acetone powder is commonly used as a source of α-glucosidase, and the mutarotase-glucose oxidase (GOD) methods commonly used to quantitate glucose produced by enzymatic hydrolysis of the substrates. In this study, we compared human Caco-2 cell extracts with rat intestinal acetone powder extracts. We also compared high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) with the mutarotase-GOD method. The sensitivity of HPAE-PAD was higher than that of mutarotase-GOD. The glucose concentration quantified by HPAE-PAD was similar to that quantified using the mutarotase-GOD method. In the maltase reaction, 1-deoxynojirimycin (1-DNJ) exerted a more potent inhibitory effect on human enzymes than on rat enzymes. This order was reversed during the sucrase reaction. These results suggested that the combined use of Caco-2 cell extracts and HPAE-PAD is advantageous for use in α-glucosidase-related basic research.

Determination of arbutin in vitro and in vivo by LC-MS/MS: Pre-clinical evaluation of natural product arbutin for its early medicinal properties.

ETHNOPHARMACOLOGICAL RELEVANCE: Arbutin is a naturally occurring glucoside extracted from plants, known for its antioxidant and tyrosinase inhibiting properties. It is widely used in cosmetic and pharmaceutical industries. With in-depth study of arbutin, its application in disease treatment is expanding, presenting promising development prospects. However, reports on the metabolic stability, plasma protein binding rate, and pharmacokinetic properties of arbutin are scarce. AIM OF THE STUDY: The aim of this study is to enrich the data of metabolic stability and pharmacokinetics of arbutin through the early pre-clinical evaluation, thereby providing some experimental basis for advancing arbutin into clinical research. MATERIALS AND METHODS: We developed an efficient and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for determining arbutin in plasma. We investigated the metabolic and pharmacokinetic properties of arbutin through in vitro metabolism assay, cytochrome enzymes P450 (CYP450) inhibition studies, plasma protein binding rate analysis, Caco-2 cell permeability tests, and rat pharmacokinetics to understand its in vivo performance. RESULTS: In vitro studies show that arbutin is stable, albeit with some species differences. It exhibits low plasma protein binding (35.35 ± 11.03% âˆ¼ 40.25 ± 2.47%), low lipophilicity, low permeability, short half-life (0.42 ± 0.30 h) and high oral bioavailability (65 ± 11.6%). Arbutin is primarily found in the liver and kidneys and is eliminated in the urine. It does not significantly inhibit CYP450 up to 10 µM, suggesting a low potential for drug interactions. Futhermore, preliminary toxicological experiments indicate arbutin's safety, supporting its potential as a therapeutic agent. CONCLUSION: This study provides a comprehensive analysis the drug metabolism and pharmacokinetics (DMPK) of arbutin, enriching our understanding of its metabolism stability and pharmacokinetics properties, It establishes a foundation for further structural optimization, pharmacological studies, and the clinical development of arbutin.

Identification of Corn Peptides with Alcohol Dehydrogenase Activating Activity Absorbed by Caco-2 Cell Monolayers.

Alcohol dehydrogenase (ADH) plays a pivotal role in constraining alcohol metabolism. Assessing the ADH-activating activity in vitro can provide insight into the capacity to accelerate ethanol metabolism in vivo. In this study, ADH-activating peptides were prepared from corn protein meal (CGM) using enzymatic hydrolysis, and these peptides were subsequently identified following simulated gastrointestinal digestion and their absorption through the Caco-2 cell monolayer membrane. The current investigation revealed that corn protein hydrolysate hydrolyzed using alcalase exhibited the highest ADH activation capability, maintaining an ADH activation rate of 52.93 ± 2.07% following simulated gastrointestinal digestion in vitro. After absorption through the Caco-2 cell monolayer membrane, ADH-activating peptides were identified. Among them, SSNCQPF, TGCPVLQ, and QPQQPW were validated to possess strong ADH activation activity, with EC50 values of 1.35 ± 0.22 mM, 2.26 ± 0.16 mM, and 2.73 ± 0.13 mM, respectively. Molecular Docking revealed that the activation of ADH occurred via the formation of a stable complex between the peptide and the active center of ADH by hydrogen bonds and hydrophobic interactions. The results of this study also suggest that corn protein hydrolysate could be a novel functional dietary element that helps protects the liver from damage caused by alcohol and aids in alcohol metabolism.

Characterization and Mechanism of a Novel Rice Protein Peptide (AHVGMSGEEPE) Calcium Chelate in Enhancing Calcium Absorption in Caco-2 Cells.

Rice protein peptides (RPP) are a potentially valuable source of high-quality calcium chelating properties. However, there is a lack of information regarding the calcium-absorption-promoting effect of RPP and its underlying mechanism. The present study adopted molecular docking methodologies to analyze the 10 most potent peptide segments from RPP. Results revealed that the peptide AHVGMSGEEPE (AHV) displayed optimal calcium binding properties (calcium-chelating capacity 55.69 ± 0.66 mg/g). Quantum chemistry analysis revealed that the AHV peptide effectively binds and forms stable complexes with calcium via the carbonyl oxygen atoms in valine at position 3 and the carbonyl of the C-terminal carboxyl group of glutamate at position 11. The spectral analysis results indicated that AHV may bind to calcium through carboxyl oxygen atoms, resulting in a transition from a smooth surface block-like structure to a dense granular structure. Furthermore, this study demonstrated that the 4 mmol/L AHV-Ca chelate (61.75 ± 13.23 µg/well) significantly increases calcium absorption compared to 1 mM CaCl2 (28.57 ± 8.59 µg/well) in the Caco-2 cell monolayer. In terms of mechanisms, the novel peptide-calcium chelate AHV-Ca derived from RPP exerts a cell-level effect by upregulating the expression of TRPV6 calcium-ion-channel-related genes and proteins (TRPV6 and Calbindin-D9k). This study provides a theoretical basis for developing functional foods with the AHV peptide as ingredients to improve calcium absorption.

Studies on the Effects of Fermentation on the Phenolic Profile and Biological Activity of Three Cultivars of Kale.

Fermentation is used not only to preserve food but also to enhance its beneficial effects on human health and achieve functional foods. This study aimed to investigate how different treatments (spontaneous fermentation or fermentation with the use of starter culture) affect phenolic content, antioxidant potential, and cholinesterase inhibitory activity in different kale cultivars: 'Halbhoner Grüner Krauser', 'Scarlet', and 'Nero di Toscana'. Chosen samples were further tested for their protective potential against the Caco-2 cell line. HPLC-MS analysis revealed that the fermentation affected the composition of polyphenolic compounds, leading to an increase in the content of rutin, kaempferol, sinapinic, and protocatechuic acids. In general, kale cultivars demonstrated various antioxidant activities, and fermentation led to an increase in total phenolic content and antioxidant activity. Fermentation boosted anti-cholinesterase activity most profoundly in 'Nero di Toscana'. Extracts of spontaneously fermented 'Scarlet' (SS) and 'Nero di Toscana' (NTS) showed cytoprotective properties, as revealed by the malondialdehyde (MDA), lactate dehydrogenase (LDH), superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) assays. Additionally, strong anti-inflammatory activity of NTS was shown by decreased release of cytokines IL-1ß and TNF-α. Collectively, the conducted studies suggest fermented kale cultivars as a potential source for functional foods.

Exploring the Hypocholesterolemic Potential of a Fucus vesiculosus Extract: Omic Insights into Molecular Mechanisms at the Intestinal Level.

High blood cholesterol levels are a major risk factor for cardiovascular diseases. A purified aqueous extract of Fucus vesiculosus, rich in phlorotannins and peptides, has been described for its potential to inhibit cholesterol biosynthesis and intestinal absorption. In this work, the effect of this extract on intestinal cells' metabolites and proteins was analysed to gain a deeper understanding of its mode of action on lipids' metabolism, particularly concerning the absorption and transport of exogenous cholesterol. Caco-2 cells, differentiated into enterocytes, were exposed to the extract, and analysed by untargeted metabolomics and proteomics. The results of the metabolomic analysis showed statistically significant differences in glutathione content of cells exposed to the extract compared to control cells, along with an increased expression of fatty acid amides in exposed cells. A proteomic analysis showed an increased expression in cells exposed to the extract compared to control cells of FAB1 and NPC1, proteins known to be involved in lipid metabolism and transport. To the extent of our knowledge, this study is the first use of untargeted metabolomics and a proteomic analysis to investigate the effects of F. vesiculosus on differentiated Caco-2 cells, offering insights into the molecular mechanism of the extract's compounds on intestinal cells.