Design of chitosan colon delivery micro/nano particles for an Achillea millefolium extract with antiproliferative activity against colorectal cancer cells.

In this study, chitosan low molecular weight (LCH) and chitosan medium molecular weight (MCH) were employed to encapsulate a yarrow extract rich in chlorogenic acid and dicaffeoylquinic acids (DCQAs) that showed antiproliferative activity against colon adenocarcinoma cells. The design of CH micro/nanoparticles to increase the extract colon delivery was carried out by using two different techniques: ionic gelation and spray drying. Ionic gelation nanoparticles obtained were smaller and presented higher yields values than spray-drying microparticles, but spray-drying microparticles showed the best performance in terms of encapsulation efficiency (EE) (> 94%), also allowing the inclusion of a higher quantity of extract. Spray-drying microparticles designed using LCH with an LCH:extract ratio of 6:1 (1.25 mg/mL) showed a mean diameter of 1.31 ± 0.21 µm and EE values > 93%, for all phenolic compounds studied. The release profile of phenolic compounds included in this formulation, at gastrointestinal pHs (2 and 7.4), showed for most of them a small initial release, followed by an increase at 1 h, with a constant release up to 3 h. Chlorogenic acid presented the higher release values at 3 h (56.91% at pH 2; 44.45% at pH 7.4). DCQAs release at 3 h ranged between 9.01- 40.73%, being higher for 1,5- and 3,4-DCQAs. After gastrointestinal digestion, 67.65% of chlorogenic and most DCQAs remained encapsulated. Therefore, spray-drying microparticles can be proposed as a promising vehicle to increase the colon delivery of yarrow phenolics compounds (mainly chlorogenic acid and DCQAs) previously described as potential agents against colorectal cancer.

Novel bioactive extract from yarrow obtained by the supercritical antisolvent-assisted technique inhibits lipid metabolism in colorectal cancer.

Background: Altered lipid metabolism in cancer is associated to dissemination and prognosis. Bioactive compounds naturally occurring in Achillea millefolium L. (yarrow) have been reported to exert antitumour activities. Food biotechnology may provide on-demand mixtures of bioactive compounds with complementary activities in cancer treatment. Methods: Supercritical-antisolvent-precipitation (SAS) has been applied to fractionate the bioactive compounds from an Ultrasound-Assisted-Extraction yarrow extract resulting in two extracts with distinct polarity, yarrow-precipitate-(PP) and yarrow-separator-(Sep). Total phenolic content and relevant essential oils have been characterized. Antioxidant, anti-inflammatory and antiproliferative activities have been compared. Moreover, the effect on the inhibition of colorectal cancer cells' bioenergetics has been evaluated. Results: Yarrow-PP exerted the highest antioxidant activity, even higher than the complete UAE-yarrow extract, meanwhile yarrow-Sep showed the highest anti-inflammatory activity, even higher than the complete UAE-yarrow extract. Interestingly, yarrow-Sep inhibited key lipid metabolic targets in CRC cells extensively shown to be implicated in cancer dissemination and prognosis -SREBF1, FASN, ABCA1 and HMGCR- and epithelial to mesenchymal targets-CDH1, ATP1B1, CDH2 and Vimentin-augmenting cell adhesion. Conclusions: In summary, SAS technology has been applied to provide a novel combination of bioactive compounds, yarrow-Sep, which merits further research to be proposed as a potential complementary nutraceutical in the treatment of CRC.

Correction: Villalva et al. Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori. Antioxidants 2022, 11, 1849.

Table S1 in the Supplementary Material (Phenolic compounds identified in yarrow samples by using HPLC-ESI-QTOF-MS) in the original paper [...].

Combination of Fenugreek and Quinoa Husk as Sources of Steroidal and Triterpenoid Saponins: Bioactivity of Their Co-Extracts and Hydrolysates.

Saponins, both steroidal and triterpenoid, exhibit distinct bioactivities. However, they are not commonly found together in natural sources; instead, sources tend to be rich in one type or another and mainly in the form of saponins rather than the sapogenin aglycones. Developing co-extracts containing both saponin or sapogenin types would be a strategy to harness their respective bioactivities, yielding multibioactive extracts. Therefore, this study evaluates the bioactivity (hypolipidemic, antioxidant, and anti-inflammatory activities) of co-extracts from fenugreek seeds (steroidal-rich saponins) and quinoa husk (triterpenoid-rich saponins), co-extracted at varying proportions, alongside their respective sapogenin-rich hydrolysates. Pancreatic lipase inhibition increased with fenugreek content in co-extracts, especially in sapogenin-rich variants. The latter substantially interfered with cholesterol bioaccessibility (90% vs. 15% in sapogenin-rich extracts). Saponin-rich co-extracts exhibited reduced cytokine release with increased fenugreek content, while sapogenin-rich counterparts showed greater reductions with higher quinoa husk content. Limited cellular antioxidant activities were observed in all extracts, with improved post-hydrolysis bioactivity. Therefore, simultaneous co-extraction of steroidal and triterpenoid sources, such as fenugreek and quinoa husk, as well as their subsequent hydrolysis, are innovative strategies for obtaining multibioactive natural extracts.

Development of Pectin Particles as a Colon-Targeted Marjoram Phenolic Compound Delivery System.

Marjoram is a culinary herb that has been widely employed in folk medicine and presents a high content in phenolics. Thus, the aim of this project was to design formulations to encapsulate phenolic compounds from marjoram to allow their release in the colon. For this purpose, pectin was used as an encapsulating agent, applying two different encapsulation techniques (ionic gelation and spray-drying), followed by a CaCl2 bath. The ionic gelation technique showed a higher yield (77%) compared to spray-drying (31%), and the particles obtained were smaller (267 nm). However, the microparticles obtained by spray-drying presented a higher encapsulation efficiency (93%). Moreover, spray-dried microparticles protected a higher percentage of the encapsulated phenolics from the action of gastrointestinal pHs and enzymes. Hence, the results showed that spray-drying was a more appropriate technique than ionic gelation for the encapsulation of marjoram phenolics in order to protect them during the gastrointestinal step, facilitating their arrival in the colon. These microparticles would also be suitable for inclusion in food matrices for the development of phenolic colon delivery systems.

Polyphenols as NLRP3 inflammasome modulators in cardiometabolic diseases: a review of in vivo studies.

The nucleotide-binding domain and leucine-rich repeat containing receptors (NLRs) are components of the innate immune system, important in coordinating the inflammatory response. Among them, NLRP3 can form inflammasomes, multiprotein complexes activating the inflammatory caspase-1 and leading, through a cell death-mediated signaling cascade, to the release of several proinflammatory cytokines. Dietary polyphenols, plant secondary metabolites, have been reported to exhibit anti-inflammatory properties, although studies have focused most on their effect on the expression of the final circulating cytokines rather than on the upstream signals activating the NLRP3 inflammasome. The present review explores current knowledge on the potential of dietary polyphenols to regulate the whole NLRP3 inflammasome pathway, in the context of cardiometabolic pathologies (obesity, cardiovascular diseases, type 2 diabetes and non-alcoholic fatty liver disease), based on in vivo studies. A clear tendency towards a decrease in the expression of the whole NLRP3 inflammasome signaling pathway when several animal models were supplemented with polyphenols was observed, commonly showing a dose-response effect; these modifications were concomitant with clinical improvements in the pathologies. Nevertheless, the diversity of doses used, the disparity in polyphenol structures tested and, particularly, the scarce clinical trials and exploration of mechanisms of action show the need to develop further research on the topic.

Reply to Franski, R.; Beszterda-Buszczak, M. Comment on "Villalva et al. Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori. Antioxidants 2022, 11, 1849".

Franski and Beszterda-Buszczak [...].

Biological Activities of Miracle Berry Supercritical Extracts as Metabolic Regulators in Chronic Diseases.

Synsepalum dulcificum (Richardella dulcifica) is a berry fruit from West Africa with the ability to convert the sour taste into a sweet taste, and for this reason, the fruit is also known as the "miracle berry" (MB). The red and bright berry is rich in terpenoids. The fruit's pulp and skin contain mainly phenolic compounds and flavonoids, which correlate with their antioxidant activity. Different polar extracts have been described to inhibit cell proliferation and transformation of cancer cell lines in vitro. In addition, MB has been shown to ameliorate insulin resistance in a preclinical model of diabetes induced by a chow diet enriched in fructose. Herein, we have compared the biological activities of three supercritical extracts obtained from the seed-a subproduct of the fruit-and one supercritical extract obtained from the pulp and the skin of MB. The four extracts have been characterized in terms of total polyphenols content. Moreover, the antioxidant, anti-inflammatory, hypo-lipidemic, and inhibition of colorectal cancer cell bioenergetics have been compared. Non-polar supercritical extracts from the seed are the ones with the highest effects on the inhibition of bioenergetic of colorectal (CRC) cancer cells. At the molecular level, the effects on cell bioenergetics seems to be related to the inhibition of main drivers of the de novo lipogenesis, such as the sterol regulatory element binding transcription factor (SREBF1) and downstream molecular targets fatty acid synthase (FASN) and stearoyl coenzyme desaturase 1 (SCD1). As metabolic reprograming is considered as one of the hallmarks of cancer, natural extracts from plants may provide complementary approaches in the treatment of cancer. Herein, for the first time, supercritical extracts from MB have been obtained, where the seed, a by-product of the fruit, seems to be rich in antitumor bioactive compounds. Based on these results, supercritical extracts from the seed merit further research to be proposed as co-adjuvants in the treatment of cancer.

Implication of the Polymeric Phenolic Fraction and Matrix Effect on the Antioxidant Activity, Bioaccessibility, and Bioavailability of Grape Stem Extracts.

The bioaccessibility and bioavailability of phenolics compounds of two grape stem extracts with different composition were studied. High polymeric extract (HPE) presented a higher content of total phenolics (TPC), procyanidins, hemicelluloses, proteins, and ashes, whereas low procyanidin extract (LPE) showed a higher fat, soluble sugars, and individual phenolic compounds content. Corresponding to its higher total phenolics content, HPE possesses a higher antioxidant activity (TEAC value). The digestion process reduced the antioxidant activity of the HPE up to 69%, due to the decrease of TPC (75%) with a significant loss of polymeric compounds. LPE antioxidant activity was stable, and TPC decreased by only 13% during the digestion process. Moreover, a higher antioxidant phenolic compounds bioavailability was shown in LPE in contrast to HPE. This behaviour was ascribed mainly to the negative interaction of polymeric fractions and the positive interaction of lipids with phenolic compounds. Therefore, this study highlights the convenience of carrying out previous studies to identify the better extraction conditions of individual bioavailable phenolic compounds with antioxidant activity, along with those constituents that could increase their bioaccessibility and bioavailability, such as lipids, although the role played by other components, such as hemicelluloses, cannot be ruled out.

Bioavailability Assessment of Yarrow Phenolic Compounds Using an In Vitro Digestion/Caco-2 Cell Model: Anti-Inflammatory Activity of Basolateral Fraction.

In this study, a combined in vitro digestion/Caco-2 model was performed with the aim to determine the phenolic compounds bioavailability of two yarrow extracts. HPLC-PAD characterisation indicated that the main components in both extracts were 3,5-dicaffeoylquinic acid (DCQA) and luteolin-7-O-glucoside. Analyses after the simulated digestion process revealed that phenolic composition was not affected during the oral phase, whereas gastric and intestinal phases represented critical steps for some individual phenolics, especially intestinal step. The transition from gastric medium to intestinal environment caused an important degradation of 3,5-DCQA (63-67% loss), whereas 3,4-DCQA and 4,5-DCQA increased significantly, suggesting an isomeric transformation within these caffeic acid derivatives. However, an approx. 90% of luteolin-7-O-glucoside was recovered after intestinal step. At the end of Caco-2 absorption experiments, casticin, diosmetin and centaureidin represented the most abundant compounds in the basolateral fraction. Moreover, this fraction presented anti-inflammatory activity since was able to inhibit the secretion of IL-1ß and IL-6 pro-inflammatory cytokines. Thus, the presence in the basolateral fraction of flavonoid-aglycones from yarrow, could be related with the observed anti-inflammatory activity from yarrow extract.

Encapsulation of Marjoram Phenolic Compounds Using Chitosan to Improve Its Colon Delivery.

In this study, chitosan particles were used to encapsulate marjoram phenolic compounds as colon-specific drug-delivery systems. The microparticles were prepared by ionic gelation and spray-drying techniques and varying amounts of polymer and extract, along with different method conditions. The spray drying of microparticles (0.75% low molecular weight chitosan dissolved in 0.4% of acetic acid) presented the best encapsulation efficiency (near 75%), with size ranges from 1.55 to 1.68 µm that allowed the encapsulation of 1.25-1.88 mg/mL of extract. Release studies of individual marjoram phenolic compounds at pH 2 and 7.4 showed that most of the compounds remained encapsulated in the microparticles. Only arbutin and vicenin II presented a high initial burst release. As the polarity of the compounds was reduced, their initial release decreased. In addition, after gastrointestinal digestion, most of marjoram phenolic compounds remained encapsulated. These results prove that chitosan microparticlescould protect the marjoram phenolic compounds during gastrointestinal digestion, specifically those related to anticancer activity, which enables their application as colon-specific delivery systems.

Case report: Functional characterization of a de novo c.145G>A p.Val49Met pathogenic variant in a case of PIGA-CDG with megacolon.

A subgroup of congenital disorders of glycosylation (CDGs) includes inherited GPI-anchor deficiencies (IGDs) that affect the biosynthesis of glycosylphosphatidylinositol (GPI) anchors, including the first reaction catalyzed by the X-linked PIGA. Here, we show the first PIGA-CDG case reported in Mexico in a male child with a moderate-to-severe phenotype characterized by neurological and gastrointestinal symptoms, including megacolon. Exome sequencing identified the hemizygous variant PIGA c.145G>A (p.Val49Met), confirmed by Sanger sequencing and characterized as de novo. The pathogenicity of this variant was characterized by flow cytometry and complementation assays in PIGA knockout (KO) cells.

Antioxidant, Anti-Inflammatory, and Antibacterial Properties of an Achillea millefolium L. Extract and Its Fractions Obtained by Supercritical Anti-Solvent Fractionation against Helicobacter pylori.

The main objective of this work is to evaluate the potential utility of an Achillea millefolium extract (yarrow extract, YE) in the control of H. pylori infection. The supercritical anti-solvent fractionation (SAF) process of YE allowed the obtaining of two different fractions: yarrow's precipitated fraction (YPF), enriched in most polar phenolic compounds (luteolin-7-O-glucoside, luteolin, and 3,5-dicaffeoylquinic acid), and yarrow's separator fraction (YSF), enriched in monoterpenes and sesquiterpenes, mainly containing camphor, artemisia ketone, and borneol. YE was effective in reducing reactive oxygen species (ROS) production in human gastric AGS cells by 16% to 29%, depending on the H. pylori strain. YPF had the highest inhibitory activity (38-40%) for ROS production. YE modulated the inflammatory response in AGS gastric cells, decreasing IL-8 production by 53% to 64%. This IL-8 inhibition also showed a strain-dependent character. YPF and YSF exhibited similar behavior, reducing IL-8 production, suggesting that both phenolic compounds and essential oils could contribute to IL-8 inhibition. YSF showed the highest antibacterial activity against H. pylori (6.3-7.1 log CFU reduction, depending on the strain) and lower MIC (0.08 mg/mL). Results obtained have shown that YE and SAF fractions (YPF and YSF) were effective as antioxidant, anti-inflammatory, and antibacterial agents regardless of the H. pylori strain characteristics.

Genetic Testing to Inform Epilepsy Treatment Management From an International Study of Clinical Practice.

Importance: It is currently unknown how often and in which ways a genetic diagnosis given to a patient with epilepsy is associated with clinical management and outcomes. Objective: To evaluate how genetic diagnoses in patients with epilepsy are associated with clinical management and outcomes. Design, Setting, and Participants: This was a retrospective cross-sectional study of patients referred for multigene panel testing between March 18, 2016, and August 3, 2020, with outcomes reported between May and November 2020. The study setting included a commercial genetic testing laboratory and multicenter clinical practices. Patients with epilepsy, regardless of sociodemographic features, who received a pathogenic/likely pathogenic (P/LP) variant were included in the study. Case report forms were completed by all health care professionals. Exposures: Genetic test results. Main Outcomes and Measures: Clinical management changes after a genetic diagnosis (ie, 1 P/LP variant in autosomal dominant and X-linked diseases; 2 P/LP variants in autosomal recessive diseases) and subsequent patient outcomes as reported by health care professionals on case report forms. Results: Among 418 patients, median (IQR) age at the time of testing was 4 (1-10) years, with an age range of 0 to 52 years, and 53.8% (n = 225) were female individuals. The mean (SD) time from a genetic test order to case report form completion was 595 (368) days (range, 27-1673 days). A genetic diagnosis was associated with changes in clinical management for 208 patients (49.8%) and usually (81.7% of the time) within 3 months of receiving the result. The most common clinical management changes were the addition of a new medication (78 [21.7%]), the initiation of medication (51 [14.2%]), the referral of a patient to a specialist (48 [13.4%]), vigilance for subclinical or extraneurological disease features (46 [12.8%]), and the cessation of a medication (42 [11.7%]). Among 167 patients with follow-up clinical information available (mean [SD] time, 584 [365] days), 125 (74.9%) reported positive outcomes, 108 (64.7%) reported reduction or elimination of seizures, 37 (22.2%) had decreases in the severity of other clinical signs, and 11 (6.6%) had reduced medication adverse effects. A few patients reported worsening of outcomes, including a decline in their condition (20 [12.0%]), increased seizure frequency (6 [3.6%]), and adverse medication effects (3 [1.8%]). No clinical management changes were reported for 178 patients (42.6%). Conclusions and Relevance: Results of this cross-sectional study suggest that genetic testing of individuals with epilepsy may be materially associated with clinical decision-making and improved patient outcomes.

Application of Pressurized Liquid Extractions to Obtain Bioactive Compounds from Tuber aestivum and Terfezia claveryi.

A PLE (pressurized liquid extraction) method was adjusted following a full-factorial experimental design to obtain bioactive-enriched fractions from Tuber aestivum and Terfezia claveryi. Temperature, time and solvent (water, ethanol and ethanol-water 1:1) parameters were investigated. The response variables investigated were: obtained yield and the levels of total carbohydrate (compounds, ß-glucans, chitin, proteins, phenolic compounds and sterols). Principal component analysis indicated water solvent and high temperatures as more adequate parameters to extract polysaccharide-rich fractions (up to 68% of content), whereas ethanol was more suitable to extract fungal sterols (up to 12.5% of content). The fractions obtained at optimal conditions (16.7 MPa, 180 °C, 30 min) were able to protect Caco2 cells from free radical exposure, acting as antioxidants, and were able to reduce secretion of pro-inflammatory cytokines in vitro: IL-6 (50%), and TNFα (80% only T. claveryi ethanol extract), as well as reduce high inhibitory activity (T. aestivum IC50: 9.44 mG/mL).

Sustainable Extraction Techniques for Obtaining Antioxidant and Anti-Inflammatory Compounds from the Lamiaceae and Asteraceae Species.

Melissa officinalis L. and Origanum majorana L., within Lamiaceae family, and Calendula officinalis L. and Achillea millefolium L., within the Asteraceae, have been considered a good source of bioactive ingredients with health benefits. In this study, the supercritical fluid extraction (SFE) using pure CO2, and the ultrasound assisted extraction (UAE) were proposed as green techniques to obtain plant-based extracts with potential antioxidant and anti-inflammatory activities. Higher values of total phenolic content and antioxidant activity were achieved in UAE ethanol:water (50:50, v/v) extracts. Meanwhile, UAE pure ethanol extracts showed greater anti-inflammatory activity. RP-HPLC-PAD-ESI-QTOF-MS/MS analysis showed a vast number of phenolic compounds in the extracts, including unreported ones. O. majorana ethanol:water extract presented the highest content of phenolics and antioxidant activity; among its composition, both rosmarinic acid and luteolin glucoside derivatives were abundant. The pure ethanol extract of A. millefolium resulted in an important content of caffeoylquinic acid derivatives, luteolin-7-O-glucoside and flavonoid aglycones, which could be related to the remarkable inhibition of TNF-α, IL-1ß and IL-6 cytokines. Besides, borneol and camphor, found in the volatile fraction of A. millefolium, could contributed to this latter activity. Thus, this study points out that O. majorana and A. millefolium are considered a promising source of bioactive ingredients with potential use in health promotion.

The Health Benefits of the Bioactive Compounds in Foods.

The health benefits of consuming certain foods have been commonly known since ancient times [...].

Valorisation of Grape Stems as a Source of Phenolic Antioxidants by Using a Sustainable Extraction Methodology.

Pressurized liquid extraction with ethanol:water mixtures was proposed for obtaining phenolic antioxidants from grape stems. The optimal extraction conditions were elucidated by using a central composite rotatable design (solvent (X1, 0-100% ethanol:water v/v), temperature (X2, 40-120 °C) and time (X3, 1-11 min)). Response surface methodology determined 30% ethanol:water, 120 °C and 10 min as the optimal extraction conditions regarding total phenolic content (TPC) (185.3 ± 2.9 mg gallic acid/g of extract) and antioxidant activity (3.55 ± 0.21 mmol Trolox/g, 1.22 ± 0.06 mmol Trolox/g and 1.48 ± 0.17 mmol Trolox/g of extract in ABTS, DPPH and ORAC methodologies, respectively). The antioxidant activity was attributed to total polymer procyanidins and flavan-3-ol monomers and oligomers, although other phenolic compound contributions should not be ruled out. Forty-two phenolic compounds were identified in the optimal extract, mainly polymer procyanidins and, to a lesser extent, monomers and oligomers of flavan-3-ols, quercetin-3-O-glucuronide, ε-viniferin, gallic and caftaric acid. Ethyl gallate, ellagic acid, protocatechuic aldehyde, delphinidin-7-O-glucoside and cyanidin-3-O-glucoside were reported for the first time in grape stem extracts. In conclusion, this study highlights the use of this winery side stream as a source of antioxidants within a sustainable food system.

Screening of bioactive compounds in truffles and evaluation of pressurized liquid extractions (PLE) to obtain fractions with biological activities.

Truffles, besides the appreciated aromatic compounds, contain other molecules with interesting bioactive properties. A screening of fungal sterols and ß-glucans within different truffle species and locations was carried out. These compounds were extracted with pressurized liquids (PLE) generating enriched fractions. Extraction efficiency was studied with a full-factorial experimental design (Response surface methodology, RSM), using water and ethanol as extraction solvents. Polysaccharides from truffle powder (TP) and the optimal PLE extract (EP) obtained were precipitated and analysed by NMR and GC-MS. THP-1 cell cultures were utilized to test immunomodulatory properties. With the optimal PLE conditions (16.7 MPa, 180 °C, 30 min) 64 and 22.5% yields were obtained respect, with water and ethanol, generating fractions containing respect, 9.1% ß-glucan and 4.5% ergosterol. NMR analyses detected (1 â†’ 3)-ß-glucan structures in truffle. The EP induced a reduction of 40% IL-1ß and 60% IL-6 pro-inflammatory cytokines secretion suggesting potential immunomodulatory activity.

Isolation and comparison of α- and ß-D-glucans from shiitake mushrooms (Lentinula edodes) with different biological activities.

A polysaccharide-enriched extract obtained from Lentinula edodes was submitted to several purification steps to separate three different D-glucans with ß-(1→6), ß-(1→3),(1→6) and α-(1→3) linkages, being characterized through GC-MS, FT-IR, NMR, SEC and colorimetric/fluorimetric determinations. Moreover, in vitro hypocholesterolemic, antitumoral, anti-inflammatory and antioxidant activities were also tested. Isolated glucans exerted HMGCR inhibitory activity, but only ß-(1→6) and ß-(1→3),(1→6) fractions showed DPPH scavenging capacity. Glucans were also able to lower IL-1ß and IL-6 secretion by LPS-activated THP-1/M cells and showed cytotoxic effect on a breast cancer cell line that was not observed on normal breast cells. These in vitro results pointed important directions for further in vivo studies, showing different effects of each chemical structure of the isolated glucans from shiitake mushrooms.