Bioprobe-RNA-seq-microRaman system for deep tracking of the live single-cell metabolic pathway chemometrics.

The integration between RNA-sequencing and micro-spectroscopic techniques has recently profiled the advanced transcriptomic discoveries on the cellular level. In the current study, by combining the sensation approach (including bio-molecules structural evaluation, high throughput next-generation sequencing (HT-NGS), and confocal Raman microscopy) the functionality on the single live cancer cells' ferroptosis and apoptosis signaling pathways is visualized. Our study reveals a hydrophobic tunnel by phycocyanin-isoprene molecule (PC-SIM) electrostatic charge within hepatoma cells (HepG2) that activates the ferritin light chain (FTL) and caspase-8 associate protein (CASP8AP2) ferroptosis responsible genes. Moreover, this research proves that PC-vanillin (VAN) stimulation induces the actin-binding factor profilin-1 (PFN1), subsequently in situ tracking its expression at 1139.75 cm-1 microRaman wavenumber. While PC-thymol (THY) induces the lysophospholipase-2 (LYPLA2) (p-value = 0.009) and acetylneuraminate-9-O-acetyltransferase (CASD1) (p-value = 0.022) at 1143.19 cm-1. Our findings establish a new concept to promote the cross-disciplinary use of instant cellular-based detection technology for intermediary evaluating the signaling cellular transcriptome.

Chinese bayberry (Myrica rubra Sieb. et Zucc.) leaves proanthocyanidins inhibit intestinal glucose transport in human Caco-2 cells.

Background: The hypoglycemic effects of Chinese bayberry leaves proanthocyanidins (BLPs) have been demonstrated. It is unclear, nevertheless, whether BLPs reduced postprandial blood glucose levels by regulating glucose uptake and glucose transport. Method: This study investigated the effect of BLPs (25, 50, and 100 µg/mL) on glucose uptake and glucose transport in human intestinal epithelial cells (Caco-2 cells). The uptake of 2-Deoxy-2-[(7-nitro-2,1,3-benzoxadiazol-4-yl) amino]-D-glucose (2-NBDG) and disaccharidases activity in Caco-2 cells were measured. The glucose transport ability across the cell membrane was determined using the established Caco-2 monolayer model. The transcript and protein levels of key glucose transporters were analyzed using real-time quantitative polymerase chain reaction (RT-qPCR) and western blotting, respectively. Results: The results showed that BLPs significantly decreased glucose uptake and disaccharidases activity (p < 0.05). Otherwise, BLPs treatment obviously inhibited glucose transport across the Caco-2 monolayer in both simulated-fast (5 mM glucose) and simulated-fed (25 mM glucose) conditions. It was attributed to the suppression of glucose transporter2 (GLUT2) and sodium-dependent glucose cotransporter 1 (SGLT1) by BLPs. BLPs were found to significantly downregulated the transcript level and protein expression of glucose transporters (p < 0.05). Meanwhile, the mRNA expression of phospholipase C (PLC) and protein kinase C (PKC) involved in the signaling pathway associated with glucose transport were decreased by BLPs. Conclusion: These results suggested that BLPs inhibited intestinal glucose transport via inhibiting the expression of glucose transporters. It indicated that BLPs could be potentially used as a functional food in the diet to modulate postprandial hyperglycemia.

Enhanced fresh-keeping capacity of printed surimi by Ca2+-nano starch-lutein and its nondestructive freshness monitoring based on 4D printed anthocyanin.

To solve undiscernible freshness changes of printed functional surimi while maintaining printed shape, 4D printable color-changing material were prepared. Firstly, based on results of printing properties and fresh-keeping of Ca2+-NS-L-surimi, it showed better printing effects (enhanced mechanical strength) and good preservation (inhibition of amino acids decomposition, bacterial growth). However, freshness changes of printed Ca2+-NS-L-surimi were not distinguished directly. To avoid that, 4D printable color-changing material-anthocyanin-hydroxypropyl methyl cellulose-xanthan gum-carrageenan (AHXK) was prepared for indicating freshness through discoloration. Printing results showed AHX with 5 % K had the most suitable mechanical strength (appropriate gel strength, texture, rheology) for printing. Based on that, AHXK had stable color (ΔE fluctuation <5) and was sensitive to pH and ammonia (obvious discoloration; ΔE > 10). Actual freshness monitoring results (co-printing of AHXK-surimi) exhibited significant discolorations, especially for HXK with 0.75 % A. It became green during refrigeration of 3-5 d (keeping fresh, ΔE < 4), brighter green at 7 d (decreased freshness, ΔE > 6), turned yellow at 9 d (spoilage, ΔE > 16), which were distinguished significantly with naked eyes rather than traditional freshness determining. In conclusion, printed AHXK-functional surimi exhibited good printing, preservation and nondestructive freshness monitoring, facilitating application of 3D printed functional surimi.

Effect of proanthocyanidins from different sources on the digestibility, physicochemical properties and structure of gelatinized maize starch.

The effect of proanthocyanidins (PAs) from Chinese bayberry leaves (BLPs), grape seeds (GSPs), peanut skins (PSPs) and pine barks (PBPs) on physicochemical properties, structure and in-vitro digestibility of gelatinized maize starch was investigated. The results showed that all PAs remarkably retarded starch digestibility, meanwhile, BLPs highlighted superiority in increasing resistant starch content from 31.29 ± 1.12 % to 68.61 ± 1.15 %. The iodine-binding affinity analysis confirmed the interaction between PAs and starch, especially the stronger binding of BLPs to amylose, which was driven by non-covalent bonds supported by XRD and FT-IR analysis. Further, we found that PAs altered the rheological properties, thermal properties and morphology structure of starch. In brief, PAs induced larger consistency, poorer flow ability, lower gelatinization temperatures and melting enthalpy change (ΔH) of starch paste. SEM and CLSM observation demonstrated that PAs facilitated starch aggregation. Our results indicated that PAs especially BLPs could be considered as potential additives to modify starch in food industry.

Protective Effect of Chinese Bayberry (Myrica rubra Sieb. et Zucc.) Pomace Wine on Oxidative Stress of Hydrogen Peroxide by Regulating Keap1/Nrf2 Pathway in HepG2 Cells.

Chinese bayberry (Myrica rubra Sieb. et Zucc.) pomace wine (CPW) is fruity and rich in bioactive compounds, with high nutritional value and antioxidant activities. This study aims to investigate the protective effect of CPW on the oxidative damage induced by hydrogen peroxide in human hepatocellular carcinoma (HepG2) cells and CPW's possible underlying mechanism. The fluorescence assay results revealed that CPW pre-treatment inhibited intracellular reactive oxygen species (ROS) accumulation in H2O2-induced HepG2 cells and cell membrane injury. Meanwhile, CPW remarkably enhanced the activity of antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), and the content of glutathione (GSH). Moreover, CPW pretreatment significantly regulated the expression of Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway-associated genes (Keap1, Nrf2, NADPH quinone oxidoreductase I (NQO1), and heme oxygenase-1 (HO-1)) and its downstream antioxidant genes (SOD, CAT, GSH, and the glutamate-cysteine ligase catalytic (GCLC) subunit) in HepG2 cells. These data demonstrated that CPW prevented H2O2-induced oxidative stress by regulating the Keap1/Nrf2 signaling pathway.

Effect of Ca2+ on 3D printing accuracy and antioxidant efficacy of nano starch-lutein surimi: mechanical properties, chemical bonds and release rate.

BACKGROUND: Nano starch-lutein (NS-L) can be used in three-dimensional (3D) printed functional surimi. However, the lutein release and printing effect are not ideal. The purpose of this study was to facilitate the function and printing properties of surimi by adding the combination of calcium ion (Ca2+ ) and NS-L. RESULTS: Printing properties, lutein release and antioxidation of printed Ca2+ -NS-L-surimi were determined. The NS-L-surimi with 20 mM kg-1 Ca2+ had the best printing effects (fine accuracy, 99 ± 1%). Compared to NS-L-surimi, the structure became denser after adding Ca2+ , the gel strength, hardness, elasticity, yield stress (τ), water holding capacity of Ca2+ -NS-L-surimi increased by about 17 ± 4%, 3 ± 1%, 9 ± 2%, 20 ± 4%, 40 ± 5% respectively. These enhanced mechanical strength and self-supporting ability to resist binding deformation and improve printing accuracy. Moreover, salt dissolution and increased hydrophobic force by Ca2+ stimulated protein stretching and aggregation, leading to enhancement of gel formation. Decreased printing effects of NS-L-surimi with excessive Ca2+ (> 20 mM kg-1 ) caused by excessive gel strength and τ, leading to strong extrusion force and low extrudability. Additionally, Ca2+ -NS-L-surimi had higher digestibility and lutein release rate (increased from 55 ± 2% to 73 ± 3%), because Ca2+ made NS-L-surimi structure porous, which promoted contact of enzyme-protein. Furthermore, weakened ionic bonds reduced electron binding bondage that combined with released lutein to provide more electrons for enhancing antioxidation. CONCLUSION: Collectively, 20 mM kg-1 Ca2+ could better promote printing process and function exertion of NS-L-surimi, facilitating the application of 3D printed functional surimi. © 2023 Society of Chemical Industry.

Novel antiosteoporotic peptides purified from protein hydrolysates of taihe black-boned silky fowl: By larval zebrafish model and molecular docking.

The black-boned silky fowl (BSF) muscle protein hydrolysate was gained by alcalase. The hydrolysate could stimulate MC3T3-E1 cell proliferation, as well as enhance alkaline phosphatas (ALP) activity and deposits of minerals. After isolation and purification, 55 peptide sequences with Mascot score over 40 were identified. Combined with molecular docking simulation and molecular dynamics analysis, two novel peptides (PASTGAAK and PGPPGTPF) were identified with the lowest binding energy of -4.99 kcal/mol and -3.07 kcal/mol with receptor BMPR1A of BMP-2/Smad pathway, showing the ability to increase BMPR1A stability. Moreover, both PASTGAAK and PGPPGTPF revealed strong anti-osteoporosis activities in the zebrafish model induced by dexamethasone. Additionally, the identified peptides could be beneficial for the differentiation of MC3T3-E1 cell for upregulating the expression of some osteoblast-related genes and proteins by stimulating BMP-2/Smad pathway. Overall, the two newly identified peptides could be the potential candidate to prevent osteoporosis.

Ultrasonic-assisted nanoencapsulation of kiwi leaves proanthocyanidins in liposome delivery system for enhanced biostability and bioavailability.

The poor biostability and bioavailability of proanthocyanidins limit their application. In this study, it was hypothesized that encapsulation in lecithin-based nanoliposomes using ultrasonic technology improves the above properties. Based on preliminary experiments, the effects of lecithin mass ratio (1-9%, wt.), pH (3.2-6.8), ultrasonic power (0-540 W), and time (0-10 min) on biostability and bioavailability of purified kiwi leaves proanthocyanidins (PKLPs) were determined. Nanoliposomes prepared optimally with lecithin (5%, wt.), pH = 3.2, ultrasonic power (270 W), and time (5 min) demonstrated a significantly (p < 0.05) improved physicochemical stability, homogeneity, and high encapsulation efficiency (73.84%) relative to control. The PKLPs bioaccessibility during in vitro digestion increased by 2.28-3.07-fold, with a remarkable sustained release and delivery to the small intestine. Similar results were obtained by in vivo analyses, showing over 200% increase in PKLPs bioaccessibility compared to the control. Thus, PKLPs-loaded nanoliposomes are promising candidates for foods and supplements for novel applications.

Improved thermal stability of roselle anthocyanin by co-pigmented with oxalic acid: Preparation, characterization and enhancement mechanism.

The enhancement effects of co-pigmentation on thermal stability of roselle anthocyanin extract (RAE) were investigated. The introduction of organic acids maintained color stability of RAE, and RAE co-pigmented with oxalic acid (OA) presented less color fading rates (19.46 ± 0.33 %) and higher redness (41.33 ± 3.51). Subsequently, suitable co-pigmentation concentration (OA:RAE = 1:2) was obtained regarding with lower ΔE (48.70 ± 2.36). Then, improvement behaviors of co-pigmentation on OA-RAE were evaluated. Results demonstrated that OA-RAE exhibited better thermal stability, as manifested by larger retention rates and more favorable thermal degradation kinetic parameters. Furthermore, both molecular docking simulation and experimental structural characterization revealed that hydrogen bonds and other non-covalent bonds made up the main parts of molecular interactions, leading to formation of stable binary complex. As a result, the aromatic ring of RAE was protected. In conclusion, the co-pigmentation of RAE via introduction of OA was effective in stability enhancement due to the generation of molecular bindings.

Ameliorative effect of bayberry leaves proanthocyanidins on high sugar diet induced Drosophila melanogaster.

Bayberry leaves proanthocyanidins (BLPs) were distributed in natural plant food, considered to have the potential for metabolic syndrome. In this study, we raised Drosophila melanogaster on high sugar diet (HSD) from the egg stage to induce hyperglycemia, and the ameliorative effect of BLPs was assessed based on this model. Phenotypical, biochemical, and molecular analyses related to diabetes mellitus pathogenesis were measured. Flies exposed to BLPs were found to suppress the HSD-induced high glucose and high triglycerides levels. Moreover, BLPs showed an inhibitory effect on carbohydrate digestive enzymes (α-amylase and α-glucosidase) activity and mRNA expression, exhibiting the potential for carbohydrate digestion retardation. Transcriptional levels of key genes associated with glycolipid metabolism were further evaluated, including dilp, InR, and downstream dAKT-dFOXO-PEPCK, together with E78, SREBP, FAS, and LSD genes, were all downregulated after BLPs-exposure, suggesting the ameliorative effect of BLPs on dysbiosis associated with the insulin signaling pathway. This study provided a new functional compound, which is beneficial to further antidiabetic therapy studies.

Evaluation of Proanthocyanidins from Kiwi Leaves (Actinidia chinensis) against Caco-2 Cells Oxidative Stress through Nrf2-ARE Signaling Pathway.

Proanthocyanidins (PAs) are considered to be effective natural byproduct and bioactive antioxidants. However, few studies have focused on their mode of action pathways. In this study, reactive oxygen species (ROS), oxidative stress indices, real-time PCR, Western blotting, confocal microscopy, and molecular docking were used to investigate the protective effect of purified kiwi leaves PAs (PKLPs) on Caco-2 cells' oxidative stress mechanisms. The results confirmed that pre-treatment with PKLPs significantly reduced H2O2-induced oxidative damage, accompanied by declining ROS levels and malondialdehyde (MDA) accumulation in the Caco-2 cells. The PKLPs upregulated the expression of antioxidative enzymes (GSH-px, CAT, T-SOD) and the relative mRNA (Nrf, HO-1, SOD-1, CAT) of the nuclear factor erythroid 2-related factor (Nrf2) signaling pathway. The protein-expressing level of the Nrf2 and its relative protein (NQO-1, HO-1, SOD-1) were significantly increased (p < 0.05) in the PKLPs pre-treatment group compared to the model group. In conclusion, the novelty of this study is that it explains how PKLPs' efficacy on the Nrf2-ARE signaling pathway, in protecting vital cells from oxidative stress, could be used for cleaner production.

Optimized Clarification Technology of Bayberry Juice by Chitosan/Sodium Alginate and Changes in Quality Characteristics during Clarification.

In this study, a novel method to clarify bayberry juice with composite clarifiers, chitosan and sodium alginate, has been designed. The optimal conditions were as follows: using chitosan 0.05 g/L first and then sodium alginate 0.05 g/L as composite clarifiers, standing for 2 h at 25 °C. The transmittance increased from 0.08 to 91.2% after treating by composite clarifiers, which was significantly higher than using chitosan (44.29%) and sodium alginate (38.46%) alone. It was also found that sedimentation time of juice treated by composite clarifiers was about 60% shorter than using single clarifiers. Meanwhile, the reduction of anthocyanin in juice was 9.16% for composite clarifiers treatment, being less than that for the single sodium alginate and previous related researches. In addition, the color and aroma of bayberry juice treated by composite clarifiers were improved. Juice treated by composite clarifiers had the highest L* value with 52.48 and looked more attractive. The present research revealed that content of beta-damascenone and dihydro-5-pentyl-2(3H)-furanone increased after treatment with composite clarifiers which contributed more to the pleasant aroma. Overall, the developed method improved the clarification effect and sensory quality, and reduced the sedimentation time, which may be promising in the production of clear bayberry juice.

Recent advances in seafood bioactive peptides and their potential for managing osteoporosis.

Marine biodiversity provides a range of diverse biological resources, including seafoods that are rich in protein and a well-balanced amino acid composition. Previous studies have shown that peptides can improve bone formation and/or inhibit bone resorption, suggesting the potential for seafood bioactive peptides (SBPs) in development of food and pharmaceuticals for management of osteoporosis. In this review, we provided an up-to-date overview of the anti-osteoporosis activity of SBPs and describe their underlying molecular mechanisms. We focus on SBPs' development, broadening the scope and depth of research, as well as strengthening in vivo and clinical research. In vitro cell cultures and in vivo animal osteoporosis models have demonstrated the potential for seafood-derived SBPs, including fish, mollusks, crustaceans, seaweed and microalgae, in preventing osteoporosis. These peptides may act by activating the signaling pathways, such as BMP/Smads, MAPK, OPG/RANKL/RANK, and NF-κB, which are associated with modulation bone health.

Ultrasound-Assisted Extraction Optimization of Proanthocyanidins from Kiwi (Actinidia chinensis) Leaves and Evaluation of Its Antioxidant Activity.

Using ultrasound (US) in proanthocyanidin (PA) extraction has become one of the important emerging technologies. It could be the next generation for studying the US mechnophore impact on the bioactive compound's functionality. The objective of this study was to demonstrate the potential of US treatment on PAs extracted from kiwifruit (Actinidia chinensis) leaves, and to provide a comprehensive chemical composition and bioactivity relationship of the purified kiwifruit leaves PAs (PKLPs). Several methods like single-factor experiments and response surface methodology (RSM) for the four affected factors on US extraction efficiency were constructed. HPLC-QTOF-MS/MS, cytotoxicity analysis, and antioxidant activity were also demonstrated. In the results, the modeling of PA affected factors showed that 40% US-amplitude, 30 mL/g dry weight (DW) solvent to solid ration (S/S), and 70 °C for 15 min were the optimum conditions for the extraction of PAs. Furthermore, PKLPs exhibited significant radical scavenging and cellular antioxidant activity (p < 0.05). In conclusion, this study's novelty comes from the broad prospects of using US in PKLP green extraction that could play an important role in maximizing this phytochemical functionality in drug discovery and food science fields.

Triacylglycerols are preferentially oxidized over free fatty acids in heated soybean oil.

In oil, free fatty acids (FFAs) are thought compared the efficiency of hydrolysis wto be the preferred substrate for lipid oxidation although triacylglycerols (TAGs) are the predominant lipid class. We determined the preferential oxidation substrate (TAGs versus FFAs) in soybean oil heated at 100 °C for 24 h, after validating a method for quantifying esterified and free lipid oxidation products (i.e., oxylipins) with mass-spectrometry. Reaction velocities and turnover (velocity per unit substrate) of FFA, and free and TAG-bound (esterified) oxylipins were determined. FFA hydrolysis rate and turnover were orders of magnitude greater (16-4217 fold) than that of esterified and free oxylipin formation. The velocity and turnover of TAG-bound oxylipins was significantly greater than free oxylipins by 282- and 3-fold, respectively. The results suggest that during heating, TAGs are preferentially oxidized over FFAs, despite the rapid hydrolysis and availability of individual FFAs as substrates for oxidation. TAG-bound oxylipins may serve as better markers of lipid oxidation.

Antiaging effects of rice protein hydrolysates on Drosophila melanogaster.

Rice protein hydrolysates (RPH) prepared by enzymatic hydrolysis have plenty of bioactive functions. Herein, we investigated the antiaging effect of RPH on Drosophila melanogaster (fruit fly) and its mechanisms. According to the results, fruit flies reared on 0.2% and 3.2% RP-supplement diet prolonged their average lifespan, 50% survival days, and the maximum lifespan, together with increasing superoxide dismutase, manganese superoxide dismutase, and catalase activity compared to those reared on basal diet. Further studies showed the lifespan extending effect of RPH was regulated by the cooperation with the intrinsic stress protection system (Nrf2/Keap1), age-related signaling pathway (TOR, S6K) and the expression of longevity genes (methuselah). In conclusion, the lifespan extending effect of RPH makes it possible to be applied in food and healthcare industry. PRACTICAL APPLICATIONS: In previous studies, rice protein hydrolysates (RPH) have been found to have strong antioxidant properties. But so far, most researches focused on the preparation, identification and in vitro antioxidant experiments of RPH, and there is still a lack of researches on its effect on the antioxidant system of fruit flies and the antiaging of fruit flies. This report showed that RPH enhanced the antioxidant system and prolonged the lifespan of Drosophila, which might help us rationally use rice peptides in functional foods.

Inhibition effect of three common proanthocyanidins from grape seeds, peanut skins and pine barks on maize starch retrogradation.

The inhibition effect of three common proanthocyanidins (PA) on the retrogradation properties of maize starch was investigated (including grape seed proanthocyanidins (GSPA), peanut skin proanthocyanidins (PSPA), and pine bark proanthocyanidins (PBPA)). Differential scanning calorimetry (DSC) and X-ray diffraction (XRD) analysis showed that PA could significantly decrease the values of melting enthalpy of retrogradation (ΔHr) and the degree of relative crystallinity, suggesting that the starch re-crystallization was retarded by PA. Scanning electron microscope (SEM) characterizations illustrated that retrograded PA-starch samples formed a looser matrix with less appearance of continuous flakes during storage. Overall, 0.5 %-2.0 % of three PAs exhibited suppression of starch retrogradation after 21-day cooling storage, mainly resulting from the PA-starch interaction. Among them, PSPA showed the most substantial inhibition effect on starch retrogradation, which might be attributed to its structural features. This study suggested that PA could be a new type of inhibitor to suppress starch retrogradation.

Copigmentation effect of three phenolic acids on color and thermal stability of Chinese bayberry anthocyanins.

The copigmentation effects of three phenolic acids (ferulic acid, sinapic acid, and syringic acid) on the color and thermal stability of Chinese bayberry anthocyanins were investigated. The three copigments of ferulic acid, sinapic acid, and syringic acid were found to have significant effects on the color enhancement of bayberry anthocyanins (p < .05). The maximum absorption wavelength of the anthocyanin aqueous solution exhibited a bathochromic shift, L* decreased, and a* increased with the increase in the molar ratio of copigments. The thermal stability of bayberry anthocyanins was significantly improved after copigmentation. No new anthocyanin derivatives appeared after copigmentation. The thermodynamic parameters (ΔG, ΔH, ΔS) of the three processes were all negative, indicating the three copigmentations were generally spontaneous and exothermic. The equilibrium constant (K) of the sinapic acid group was the greatest among the three phenolic acids, indicating that sinapic acid was more effective in stabilizing the anthocyanins.

In vitro inhibitory effects of Chinese bayberry (Myrica rubra Sieb. et Zucc.) leaves proanthocyanidins on pancreatic α-amylase and their interaction.

Chinese bayberry leaves proanthocyanidins (BLPs) belongs to the prodelphinidin category with potent EGCG unit, whose inhibition effect on α-amylase and their interaction were investigated by in vitro digestion and enzyme kinetic analysis, multi fluorescence spectroscopies (fluorescence quenching, synchronous fluorescence, and three-dimensional fluorescence), circular dichroism spectra, Fourier transform infrared spectroscopy and in silico modelling. The results revealed that BLPs was a mixed inhibitor to α-amylase with the IC50 value of 3.075 ± 0.073 µg/mL. BLPs could lead to a static fluorescence quenching of α-amylase, mainly by means of interacting with amino acids (mainly Try and Tyr residues) in one site on α-amylase molecule under the action of hydrogen bonding and/or Van der Waals force. This interaction further induced the change of secondary conformational structure, functional group structure and hydrophobicity of α-amylase, thus resulting in lowering activity. Molecular docking simulated that this binding occurred in a cavity on the surface of the α-amylase molecule, and BLPs trimer showed a relatively high binding energy. The present study provided a new insight of BLPs as an α-amylase inhibitor, which could be considered in anti-diabetic therapy.