Bioaccessibility and bioavailability of essential and potentially toxic trace elements in potato cultivars: A comprehensive nutritional evaluation.

Among the most consumed foods in the world is potato, which occupies the first place as a non-grain commodity, demonstrating the importance of its assessment concerning the population's food safety. In this study, the nutrients Ca, Mg, K, P, Cu, Mn, Fe, and Zn and the potentially toxic trace elements Cd, Cr, and Pb were evaluated considering their total contents, bioaccessible and bioavailable fractions in different potato cultivars, in an unpublished approach in the literature. The in vitro standard gastrointestinal digestion method (INFOGEST) and a model of the intestinal epithelial barrier using the Caco-2 cell line were applied for investigate the presence of metals in potato. For the macroelements, the bioaccessibility (% w/w) varied in the ranges: K (57-72 %), P (59-76 %), Mg (83-103 %), and Ca (30-123 %), whereas for the microelements were: Cu (27-74 %) and Mn (4.22-12.02, 60-119 %). The potentially of trace toxic elements, Cd and Pb, were found in 75 % of the samples, however, all the concentration values were below the maximum levels allowed of 0.10 µg/g. Chromium was determined only in potato peels and has no maximum established level. The bioaccessible and bioavailable fractions of Cd, Cr, and Pb were below the limits of quantification of the spectrometric methods (LOQ - µg/L: 0.063 Cd, 0.65 Cr, and 0.44 Pb). The potato samples were considered safe for consumption regarding the presence of potentially toxic trace elements, with a remarkable nutritional contribution.

Enantioselective analysis of the pesticide imazamox after in vitro permeability study in Caco-2 cells.

Imazamox (IMX), a chiral herbicide used in cereals and oilseed crops to control weeds, is commonly sold as a racemic mixture. Its enantiomers, being chiral compounds, may exhibit unique properties when exposed to chiral environments. While IMX enantiomers have been reported to degrade differently in soil and be toxic to some species, their effects on human systems remain poorly understood. This study utilized Caco-2 (human colon adenocarcinoma cell line) cells to assess the in vitro permeability of a racemic mixture of IMX and its isolated enantiomers. Additionally, the study aimed to evaluate whether the metabolite imazamox-O-desmethyl (IMX-D) forms during the permeability process. An enantioselective chromatographic method was developed, fully validated, and the apparent permeability values were obtained. The apparent permeability of rac-IMX, (+)-IMX, and (-)-IMX was determined to be 4.15 × 10-5, 5.78 × 10-5, and 7.33 × 10-5 cm s-1, respectively. These findings suggest that IMX exhibits high intestinal permeability, with an enantioselective absorption for (-)-IMX as compared to (+)-IMX. Finally, the permeability study in Caco-2 cells revealed that the metabolite IMX-D was not generated.

Effects of Branched-Chain Amino Acids on the Inflammatory Response Induced by LPS in Caco-2 Cells.

Branched-chain amino acids (BCAA) are essential for maintaining intestinal mucosal integrity. However, only a few studies have explored the role of BCAA in the modulation of intestinal inflammation. In this study, we investigated in vitro effects of BCAA on the inflammatory response induced by lipopolysaccharide (LPS) (1 µg/mL) in Caco-2 cells. Caco-2 cells were assigned to six groups: control without BCAA (CTL0), normal BCAA (CTL; 0.8 mM leucine, 0.8 mM isoleucine, and 0.8 mM valine); leucine (LEU; 2 mM leucine), isoleucine (ISO; 2 mM isoleucine), valine (VAL; 2 mM valine), and high BCAA (LIV; 2 mM leucine, 2 mM isoleucine, and 2 mM valine). BCAA was added to the culture medium 24 h before LPS stimulation. Our results indicated that BCAA supplementation did not impair cell viability. The amino acids leucine and isoleucine attenuated the synthesis of IL-8 and JNK and NF-kB phosphorylation induced by LPS. Furthermore, neither BCAA supplementation nor LPS treatment modulated the activity of glutathione peroxidase or the intracellular reduced glutathione/oxidized glutathione ratio. Therefore, leucine and isoleucine exert anti-inflammatory effects in Caco-2 cells exposed to LPS by modulating JNK and NF-kB phosphorylation and IL-8 production. Further in vivo studies are required to validate these findings and gather valuable information for potential therapeutic or dietary interventions.

Evaluation of Virulence of Salmonella Infantis and Salmonella Enteritidis with In Vitro and In Vivo Models.

Salmonella Infantis and Enteritidis serovars have been reported as important causes of salmonellosis in humans worldwide. However, the virulence of these two serovars has yet to be compared. To evaluate the virulence of Salmonella Infantis (n = 23) and Salmonella Enteritidis (n = 7), we used two models: the Caco2 cells model (in vitro) and the Galleria mellonella model (in vivo). Additionally, the virulence genes of all tested strains were contrasted with phenotypic outcomes. Results showed that adhesion means were 18.2% for Salmonella Enteritidis and 38.2% for Salmonella Infantis strains. Invasion means were 77.1% for Salmonella Enteritidis and 56.2% for Salmonella Infantis strains. Significant differences were found between serovars in adherence and invasion assays. Mortality rates (58% for Salmonella Enteritidis and 62.6% for Salmonella Infantis) were not significantly different between serotypes. The distribution of virulence genes showed that genes fae (fimbrial adherence determinants) and shdA (nonfimbrial adherence determinants) were only found in Salmonella Infantis strains. On the other hand, the rck gene (invasion) and Plasmid-encoded fimbriae genes (pef A, B, C, D) were present in Salmonella Enteritidis exclusively. In conclusion, this study shows that Salmonella Enteritidis has a higher virulence potential under experimental conditions than Salmonella Infantis. However, more studies are needed to determine the risk that Salmonella Infantis could represent compared with Salmonella Enteritidis. Moreover, other in vivo models should be considered to assess the virulence of these serovars.

Transglycosylation catalysed by Caco-2 membrane disaccharidases: A new approach to understand carbohydrates digestibility.

Under appropriate experimental conditions, some glycoside hydrolases can catalyze transglycosylation reactions; a hypothesis associated with this is that the glycosidic linkages formed will be preferentially hydrolyzed under optimal conditions. Therefore, the hydrolytic and transglycosylation activities of isolated membranes from differentiated Caco-2 cells on sucrose, maltose and isomaltulose were evaluated. After the enzymatic reactions, the di- and trisaccharides obtained were identified by gas chromatography coupled to a mass spectrometer. Differentiated Caco-2 cell membranes exerted hydrolytic and transglycosylation activities towards the studied disaccharides. The obtained di- and trisaccharides were detected for the first time using human cell models. Due to the absence of maltase-glucoamylase complex (MGAM) in Caco-2 cells, and the known hydrolytic activity of sucrase-isomaltase (SI) towards sucrose, maltose and isomaltulose, it is plausible that the glycosidic linkages obtained after the transglycosylation reaction, mainly α-glucosyl-fructoses and α-glucosyl-glucoses, were carried out by SI complex. This approach can be used as a model to explain carbohydrate digestibility in the small intestine and as a tool to design new oligosaccharides with low intestinal digestibility.

Improved functional properties of the potential probiotic Lacticaseibacillus paracasei ItalPN16 growing in cheese whey.

The production of probiotic bacteria requires specific and expensive culture media for maintain their viability and metabolic response during gastro-intestinal transit and cell adhesion process. The aim of this study was to compare the ability of the potential probiotic Laticaseibacillus paracasei ItalPN16 to grow in plain sweet whey (SW) and acid whey (AW), evaluating changes in some probiotic properties related to the culture media. Pasteurized SW and AW were suitable media for L. paracasei growth, since counts above 9 Log CFU/ml were achieved using <50% of the total sugars in both whey samples after 48 h at 37°C. The L. paracasei cells obtained from AW or SW cultures showed increased resistance to pH 2.5 and 3.5, higher autoaggregation, and lower cell hydrophobicity, as compared with the control of MRS. SW also improved the biofilm formation ability and cell adhesion capability to Caco-2 cells. Our results indicate that the L. paracasei adaptation to the SW conditions, inducing metabolic changes that improved its stability to acid stress, biofilm formation, autoaggregation, and cell adhesion properties, which are important functional probiotic properties. Overall, the SW could be considered as low-cost culture medium for sustainable biomass production of L. paracasei ItalPN16.

Cocoa powder and fermented jackfruit seed flour: A comparative cell-based study on their potential antioxidant and anti-inflammatory activities after simulated gastrointestinal digestion.

BACKGROUND: Jackfruit seed flour can be used as a cocoa aroma replacer with similar technological properties. The purpose of this study was to investigate the in vivo toxicity and in vitro antioxidant activity of fermented jackfruit seed flour (Fjs) and non-alkaline cocoa powder (Nac). RESULTS: Fjs and Nac extracts (Fjs-E and Nac-E) were produced and submitted to in vitro gastrointestinal digestion producing digested fractions named Fjs-D and Nac-D, respectively. Nac-E showed over two-fold higher oxygen radical absorbance capacity (ORAC) than Fjs-E. However, after simulated gastrointestinal digestion (in vitro), there were no significant differences between Nac-D and Fjs-D (P < 0.01). Similarly, the cellular antioxidant activity (CAA) of Nac-D and Fjs-D was not significantly different (P < 0.01). The anti-inflammatory assay in transgenic RAW 264.7 murine macrophages showed that Fjs-E did not affect cell viability up to 300 µg mL-1 (P > 0.05) and reduced by 15% the release of TNF-α (P < 0.05). Fjs-D did not affect cell viability up to 300 µg mL-1 (P > 0.05) and showed 58% reduction of NF-κB activation (P < 0.05), with no effects on TNF-α levels. Treatment with Nac-E up to 300 µg mL-1 did not decrease cell viability (P > 0.05) and reduced the release of TNF-α levels by 34% and 66% at 100 and 300 µg mL-1 , respectively (P < 0.05). Nac-D did not reduce the NF-κB activation or TNF-α levels at any tested concentration. CONCLUSION: Collectively, these findings indicate that Fjs is a safe and promising functional ingredient with biological activities even after gastrointestinal digestion. © 2023 Society of Chemical Industry.

Guidance for formulating ingredients/products from Chlorella vulgaris and Arthrospira platensis considering carotenoid and chlorophyll bioaccessibility and cellular uptake.

This study aimed to investigate the impact of different microalgal matrices on the bioaccessibility and uptake by Caco-2 cells of carotenoids and chlorophylls. In this way, the microalgal ingredients/products (whole dry biomass [WDB], whole ultrasonicated paste [WUP], and liposoluble pigment emulsion [LPE]) obtained from Chlorella vulgaris and Arthrospira platensis were submitted to in vitro simulated digestion. Apical uptake of pigments in micelles generated during the simulated digestion by Caco-2 human intestinal cells was determined. The influence of simulated digestion on carotenoid and chlorophyll stability and bioaccessibility was assessed by HPLC-PDA-MS/MS and the carotenoids and chlorophylls' bioaccessibility and cellular uptake were shown to be boosted according to the matrix (LPE > WUP > WDB). Our findings showed that Chlorella vulgaris and Arthrospira platensis could be considered in formulations when carotenoids and chlorophylls are the target molecules in the ingredients/products.

Formulations with microencapsulated Fe-peptides improve in vitro bioaccessibility and bioavailability.

The bioaccessibility and the bioavailability of iron complexed to peptides (active) in microparticles forms contained in dry beverages formulations were evaluated. The peptide-iron complexes microparticles were obtained by spray drying and added in three dry formulations (tangerine, strawberry, and chocolate flavors). The peptides isolated by iron ion affinity (IMAC-Fe III) had their biological activity predicted by BIOPEP® database and were evaluated by molecular coupling. The bioaccessibility was evaluated by solubility and dialysability and the bioavalability was assessed by Caco-2 cellular model. The proportion 10:1 of peptide-iron complexes presented higher rates of bioaccessibility (49%) and bioavailability (56%). The microparticle with peptide-iron complex showed greater solubility after digestion (39.1%), bioaccessibility (19.8%), and bioavailability (34.8%) than the ferrous sulfate salt (control) for the three assays (10.2%; 12.9%; 9.7%, respectively). Tangerine and strawberry formulations contributed to the iron absorption according to the results of bioaccessibility (36.2%, 30.0% respectively) and bioavailability (80.5%, 84.1%, respectively). The results showed that iron peptide complexation and microencapsulation process improve the bioaccessibility and bioavailability when incorporated into formulations.

Bacteriostatic effect of Echeveria extracts on diarrheagenic E. coli pathotypes and non-cytotoxicity on human Caco-2 cells.

INTRODUCTION: Diarrheagenic Escherichia coli pathotypes are important aetiological agents of diarrhoeal illness among children from less developed areas, worldwide. Diarrheagenic E. coli pathotypes strains are increasingly becoming drug resistant, thus effective and accessible therapeutic alternatives are required for their treatment; herbal extracts may be a potential alternative. AIMS: to evaluate Echeveria craigiana, E. kimnachii, and E. subrigida methanol extracts antibacterial effect on six diarrheagenic E. coli reference strains and on human colorectal adenocarcinoma cells viability and cytokine production. METHODOLOGY: Diarrheagenic E. coli pathotypes reference strains: typical enteropathogenic E2348/69, enterotoxigenic H10407, enterohaemorrhagic O157:H7/EDL933, enteroinvasive E11, diffusely adherent C18451-A, and enteroaggregative 042 E. coli. E craigiana, E. kimnachii, and E. subrigida leaves, collected at Sinaloa, Mexico, were freeze-dried and macerated in methanol solvent. Antibacterial activity was determined by a novel method developed in our laboratory, bacterial oxygen consumption by polarographic oxygen electrode technique and membrane integrity by two methods (live/dead and protein leakage assays). Colorectal adenocarcinoma cells viability by MTT assay and cytokine production using a Cytometric Bead Array kit. RESULTS: Extracts concentrations of 100 µg/mL and 5-hour incubation, reduced more than 93% the growth of all diarrheagenic E. coli pathotypes tested strains and significantly decreased bacterial oxygen consumption, like bacteriostatic antibiotics. After 24-hour incubation methanol extracts had a differential antibacterial effect on each diarrheagenic E. coli pathotypes strain. Echeveria extracts did not have any effect on viability and cytokine production of colorectal adenocarcinoma cells. CONCLUSIONS: Echeveria methanol extracts have a bacteriostatic effect on all diarrheagenic E. coli pathotypes strains, thus potentially they could be used as antibacterial agents on diarrheagenic E. coli pathotypes-contaminated products and on patients with diarrheagenic E. coli pathotypes infections.

Toxicological assessment of a bioactive extract from Triplaris gardneriana Wedd. seeds using alternative models.

The Triplaris gardneriana Wedd. seeds extract has great therapeutic potential due to numerous biological activities such as antioxidant, antibacterial and anti-inflammatory, which are associated with phenolic content. Although this herbal preparation has shown many benefits, recently their toxicity profile has begun to be explored. In this present study, the toxic effects of T. gardneriana seeds ethanolic extract (EETg) on biological systems of different taxonomical groups and levels of complexity (from cell culture to lower vertebrates) were assessed, through a variety of viability and toxicological assays. It was found that EETg did not impair the Saccharomyces cerevisiae growth at the highest tested concentration (200 µg/mL), and no toxicant evidence was observed in Aedes aegypti larvae or in Drosophila melanogaster adult stage. Contrarily, the extract reduced the viability of undifferentiated Caco-2 cells (250 µg/mL, 40% of viable cells), but did not affect differentiated ones. The embryotoxicity in Danio rerio model showed a LC50 of 7.41 mg/L (95% confidence interval, 4.78 - 11.49 mg/L). EETg did not show signs of toxicity in the majority of the models used, but lethality and malformations in zebrafish embryos occurred. Further analyses are needed to better understand the selective toxicity mechanism of EETg on zebrafish, as well as whether the toxic effects happen in higher vertebrates.

Semysinthetic biflavonoid Morelloflavone-7,4',7″,3‴,4‴-penta-O-butanoyl is a more potent inhibitor of Proprotein Convertases Subtilisin/Kexin PC1/3 than Kex2 and Furin.

BACKGROUND: Garcinia brasiliensis is a species native to the Amazon forest. The white mucilaginous pulp is used in folk medicine as a wound healing agent and for peptic ulcer, urinary, and tumor disease treatments. The activity of the proprotein convertases (PCs) Subtilisin/Kex is associated with the development of viral, bacterial and fungal infections, osteoporosis, hyperglycemia, atherosclerosis, cardiovascular, neurodegenerative and neoplastic diseases. METHODS: Morelloflavone (BF1) and semisynthetic biflavonoid (BF2, 3 and 4) from Garcinia brasiliensis were tested as inhibitor of PCs Kex2, PC1/3 and Furin, and determined IC50, Ki, human proinflammatory cytokines secretion in Caco-2 cells, mechanism of inhibition, and performed molecular docking studies. RESULTS: Biflavonoids were more effective in the inhibition of neuroendocrine PC1/3 than mammalian Furin and fungal Kex2. BF1 presented a mixed inhibition mechanism for Kex2 and PC1, and competitive inhibition for Furin. BF4 has no good interaction with Kex2 and Furin since carboxypropyl groups results in steric hindrance to ligand-protein interactions. Carboxypropyl groups of BF4 promote steric hindrance with Kex2 and Furin, but effective in the affinity of PC1/3. BF4 was more efficient at inhibiting PCl/3 (IC50 = 1.13 µM and Ki = 0,59 µM, simple linear competitive mechanism of inhibition) than Kex2, Furin. Also, our results strongly suggested that BF4 also inhibits the endogenous cellular PC1/3 activity in Caco-2 cells, since PC1/3 inhibition by BF4 causes a large increase in IL-8 and IL-1ß secretion in Caco-2 cells. CONCLUSIONS: BF4 is a potent and selective inhibitor of PC1/3. GENERAL SIGNIFICANCE: BF4 is the best candidate for further clinical studies on inhibition of PC1/3.

Heat-Labile Toxin from Enterotoxigenic Escherichia coli Causes Systemic Impairment in Zebrafish Model.

Heat-labile toxin I (LT-I), produced by strains of enterotoxigenic Escherichia coli (ETEC), causes profuse watery diarrhea in humans. Different in vitro and in vivo models have already elucidated the mechanism of action of this toxin; however, their use does not always allow for more specific studies on how the LT-I toxin acts in systemic tracts and intestinal cell lines. In the present work, zebrafish (Danio rerio) and human intestinal cells (Caco-2) were used as models to study the toxin LT-I. Caco-2 cells were used, in the 62nd passage, at different cell concentrations. LT-I was conjugated to FITC to visualize its transport in cells, as well as microinjected into the caudal vein of zebrafish larvae, in order to investigate its effects on survival, systemic traffic, and morphological formation. The internalization of LT-I was visualized in 3 × 104 Caco-2 cells, being associated with the cell membrane and nucleus. The systemic traffic of LT-I in zebrafish larvae showed its presence in the cardiac cavity, yolk, and regions of the intestine, as demonstrated by cardiac edema (100%), the absence of a swimming bladder (100%), and yolk edema (80%), in addition to growth limitation in the larvae, compared to the control group. There was a reduction in heart rate during the assessment of larval survival kinetics, demonstrating the cardiotoxic effect of LT-I. Thus, in this study, we provide essential new depictions of the features of LT-I.

Bioaccessibility and Cellular Uptake of Carotenoids Extracted from Bactris gasipaes Fruit: Differences between Conventional and Ionic Liquid-Mediated Extraction.

Currently, on an industrial scale, synthetic colorants are used in many fields, as well as those extracted with conventional organic solvents (COSs), leading to several environmental issues. Therefore, we developed a sustainable extraction and purification method mediated by ionic liquids (IL), which is considered an alternative high-performance replacement for COSs. Carotenoids are natural pigments with low bioaccessibility (BCT) and bioavailability (BV) but with huge importance to health. To investigate if the BCT and cellular uptake of the carotenoids are modified by the extraction method, we conducted a comparison assay between both extraction procedures (IL vs. COS). For this, we used the Amazonian fruit Bactris gasipaes, a rich source of pro-vitamin A carotenoids, to obtain the extract, which was emulsified and subjected to an in vitro digestion model followed by the Caco-2 cell absorption assay. The bioaccessibility of carotenoids using IL was better than those using COS (33.25%, and 26.84%, respectively). The cellular uptake of the carotenoids extracted with IL was 1.4-fold higher than those extracted using COS. Thus, IL may be a feasible alternative as extraction solvent in the food industry, replacing COS, since, in this study, no IL was present in the final extract.

Insights on the intestinal absorption of chlorophyll series from microalgae.

The bioaccessibility and subsequent uptake by Caco-2 human intestinal cells of chlorophyll pigments from Scenedesmus obliquus were determined for the first time. In order to evaluate the impact of different types of the matrix on bioaccessibility of chlorophyll from microalgae, three different products were evaluated: isolated chlorophyll extract (ICE); wet ultrasonicated biomass (WUB); and whole dried biomass (WDB). The samples were submitted to in vitro digestion model according to the INFOGEST protocol, and Caco-2 cells determined the intestinal uptake. Chlorophyll pigments were determined by HPLC-PDA-MS/MS. A total of ten chlorophyll pigments (8,318.48 µg g-1) were separated in S. obliquus biomass, with chlorophyll a (3,507.76 µg g-1) and pheophytin a' (1,598.09 µg g-1) the major ones. After in vitro digestion, all tested products showed bioaccessible chlorophylls. However, the total bioaccessibility results were as follows: ICE (33.45%), WUB (2.65%), WDB (0.33%). Five compounds were bioaccessible in ICE, three in WUB, and one in WDB. The hydroxypheophytin a showed the highest bioaccessibility (212%) in ICE, while pheophytin a' in WUB (11%) and WDB (2%). As a result, bioavailability estimates of ICE using the Caco-2 cell showed hydroxypheophytin a (102.53%), followed by pheophytin a' (64.69%) as the chlorophyll pigments most abundant in intestinal cells. In summary, from a nutritional perspective, these three types of the matrix (WDB, WUB, and ICE) influence the promotion of chlorophyll bioaccessibility. In this way, the data suggest that chlorophylls bioaccessibility from ICE is greater than that in WDB and WUB. Therefore, ICE should be considered a product that provides bioavailable chlorophyll and could be the best choice, such as ingredients in the development of functional foods chlorophyll-based.

Heat-Labile toxin from enterotoxigenic Escherichia coli causes systemic impairment in Zebrafish model

Heat-labile toxin I (LT-I), produced by strains of enterotoxigenic Escherichia coli (ETEC), causes profuse watery diarrhea in humans. Different in vitro and in vivo models have already elucidated the mechanism of action of this toxin; however, their use does not always allow for more specific studies on how the LT-I toxin acts in systemic tracts and intestinal cell lines. In the present work, zebrafish (Danio rerio) and human intestinal cells (Caco-2) were used as models to study the toxin LT-I. Caco-2 cells were used, in the 62nd passage, at different cell concentrations. LT-I was conjugated to FITC to visualize its transport in cells, as well as microinjected into the caudal vein of zebrafish larvae, in order to investigate its effects on survival, systemic traffic, and morphological formation. The internalization of LT-I was visualized in 3 × 104 Caco-2 cells, being associated with the cell membrane and nucleus. The systemic traffic of LT-I in zebrafish larvae showed its presence in the cardiac cavity, yolk, and regions of the intestine, as demonstrated by cardiac edema (100%), the absence of a swimming bladder (100%), and yolk edema (80%), in addition to growth limitation in the larvae, compared to the control group. There was a reduction in heart rate during the assessment of larval survival kinetics, demonstrating the cardiotoxic effect of LT-I. Thus, in this study, we provide essential new depictions of the features of LT-I.

Determination of l-(+)-bornesitol, the hypotensive constituent of Hancornia speciosa, in rat plasma by LC-MS/MS and its application on a pharmacokinetic study.

Hancornia speciosa is a medicinal plant with proven antihypertensive activity. The cyclitol l-(+)-bornesitol is the main constituent of its leaves and is a potent inhibitor of the angiotensin-converting enzyme. We herein investigated the pharmacokinetic properties of bornesitol administered orally to Wistar rats, as well as bornesitol permeation in Caco-2 cells. Bornesitol was isolated and purified from an ethanol extract of H. speciosa leaves. An ultra-high performance liquid chromatography coupled with electrospray ionization mass spectrometry (UPLC-ESI-MS/MS) method was developed and validated to quantify bornesitol in rat plasma based on Multiple Reaction Monitoring, using pentaerythritol as an internal standard. Pharmacokinetics was evaluated by the administration of single doses via intravenous in bolus (3 mg/kg) and gavage (3, 15 and 25 mg/kg). Bornesitol permeation was assayed in a transwell Caco-2 cells model, tested alone, or combined with rutin, or as a constituent of H. speciosa extract, using a developed and validated UPLC-ESI-MS/MS method. All assayed validation parameters (selectivity, residual effect, matrix effect, linearity, precision, accuracy and stability of analyte in plasma and solution) for the bioanalytical method met the acceptance criteria established by regulatory guidelines. Bornestiol reached peak plasma concentration within approximately 60 min after oral administration with a half-life ranging from 72.15 min to 123.69 min. The peak concentration and area under the concentration-time curve of bornesitol did not rise proportionally with the increasing doses, suggesting a non-linear pharmacokinetics in rats and the oral bioavailability ranged from 28.5%-59.3%. Bornesitol showed low permeability in Caco-2 cells, but the permeability apparently increased when it was administered either combined with rutin or as a constituent of H. speciosa extract. In conclusion, bornesitol was rapidly absorbed after a single oral administration to rats and followed a non-linear pharmacokinetics. The obtained data will be useful to guide further pre-clinical development of bornesitol-containing herbal preparations of H. speciosa as an antihypertensive agent.

Optimization of the Red Tilapia (Oreochromis spp.) Viscera Hydrolysis for Obtaining Iron-Binding Peptides and Evaluation of In Vitro Iron Bioavailability.

Iron deficiencies continue to cause significant health problems in vulnerable populations. A good strategy to combat mineral deficiency includes fortification with iron-binding peptides. This research aims to determine the optimal conditions to hydrolyze red tilapia viscera (RTV) using Alcalase 2.4 L and recovery of iron-binding protein hydrolysate. The result showed that under the optimal hydrolysis condition including pH 10, 60 °C, E/S ratio of 0.306 U/g protein, and substrate concentration of 8 g protein/L, the obtained hydrolysate with 42.5% degree of hydrolysis (RTVH-B), displayed the maximal iron-binding capacity of 67.1 ± 1.9%. Peptide fractionation was performed using ultrafiltration and the <1 kDa fraction (FRTVH-V) expressed the highest iron-binding capacity of 95.8 ± 1.5%. Iron content of RTVH-B and its fraction was assessed, whereas iron uptake was measured indirectly as ferritin synthesis in a Caco-2 cell model and the result showed that bioavailability of bound minerals from protein complexes was significantly higher (p < 0.05) than iron salt in its free form, increased 4.7 times for the Fe2+-RTVH-B complex. This research suggests a potential application of RTVH-B as dietary supplements to improve iron absorption.

Matrix effects of the hydroethanolic extract and the butanol fraction of calyces from Physalis peruviana L. on the biopharmaceutics classification of rutin.

OBJECTIVES: The Biopharmaceutics Classification System (BCS) categorizes active pharmaceutical ingredients according to their solubility and permeability properties, which are susceptible to matrix or formulation effects. The aim of this research was to evaluate the matrix effects of a hydroethanolic extract of calyces from Physalis peruviana L. (HEE) and its butanol fraction (BF), on the biopharmaceutics classification of their major compound, quercetin-3-O-rutinoside (rutin, RU). METHODS: Rutin was quantified by HPLC-UV, and Caco-2 cell monolayer transport studies were performed to obtain the apparent permeability values (Papp ). Aqueous solubility was determined at pH 6.8 and 7.4. KEY FINDINGS: The Papp values followed this order: BF > HEE > RU (1.77 ± 0.02 > 1.53 ± 0.07 > 0.90 ± 0.03 × 10-5  cm/s). The lowest solubility values followed this order: HEE > RU > BF (2.988 ± 0.07 > 0.205 ± 0.002 > 0.189 ± 0.005 mg/ml). CONCLUSIONS: According to these results, rutin could be classified as BCS classes III (high solubility/low permeability) and IV (low solubility/low permeability), depending on the plant matrix. Further work needs to be done in order to establish how apply the BCS for research and development of new botanical drugs or for bioequivalence purposes.

Oral nanoparticles based on gellan gum/pectin for colon-targeted delivery of resveratrol.

Nanoparticles based on gellan gum/pectin blends were designed for colon-targeted release of resveratrol (RES). Their impact on drug release rates and permeability were evaluated using Caco-2 cell model and mucus secreting triple co-culture model. Polymeric nanoparticles (PNP) were successfully prepared by nebulization/ionotropic gelation, achieving high drug loading (>80%). PNP were spherical with a low positive charge density (+5mV) and exhibited diameters of around 330 nm. Developed PNP were able to promote effective modulation of drug release rates, so that only 3% of RES was released in acidic media over 2 h, and, in pH 6.8, the drug was released in a sustained manner, reaching 85% in 30 h. The permeability of RES-loaded PNP in the Caco-2 model was 0.15%, while in the triple co-culture model, in the presence of mucus, it reached 5.5%. The everted gut sac experiment corroborated the low permeability of RES-loaded PNP in the presence or absence of mucus and highlighted their high ability to interact with the intestinal tissue. Results indicate that the novel PNP developed in this work are safe and promising carriers for controlled delivery of RES at the colon.