Ethanol Precipitation Research Articles

The impact of extraction processes on the physicochemical, functional properties and structures of bamboo shoot protein

This study aimed to extract bamboo shoot protein (BSP) using different extraction approaches and compare their functional and physicochemical properties with commercial protein ingredients, including whey protein and soy protein isolates. The extraction methods including alkali extraction (AE), salt extraction (SE), and phosphate-aided ethanol precipitation (PE) were used. An enhanced solvent extraction method was utilized in combination, resulting in a significant improvement in the protein purity, which reached 81.59 %, 87.36 %, and 67.08 % respectively. The extraction methods had significant effects on the amino acid composition, molecular weight distribution, and functional properties of the proteins. SE exhibited the best solubility and emulsification properties. Its solubility reached up to 93.38 % under alkaline conditions, and the emulsion stabilized by SE with enhanced solvent extraction retained 60.95 % stability after 120 min, which could be attributed to its higher protein content, higher surface hydrophobicity, and relative more stable and organized protein structure. All three BSP samples demonstrated better oil holding capacity, while the SE sample showed comparable functional properties to soy protein such as foaming and emulsifying properties. These findings indicate the potential of BSP as an alternative plant protein ingredient in the food industry.

Structural characterization and immunostimulant activities of polysaccharides fractionated by gradient ethanol precipitation method from Panax ginseng C. A. Meyer.

Panax ginseng C. A. Meyer is a dual-purpose plant for medicine and food, its polysaccharide is considered as an immune enhancer. Four polysaccharides, WGP-20-F, WGP-40-F, WGP-60-F and WGP-80-F were obtained from ginseng via water extraction and gradient ethanol precipitation with different molecular weights (Mw) of 1.720 × 106, 1.434 × 106, 4.225 × 104 and 1.520 × 104Da, respectively. WGP-20-F and WGP-40-F which with higher Mw and a triple-helix structure are glucans composed of 4-ɑ-Glcp, do not show remarkable immunoregulatory effects. WGP-60-F and WGP-80-F are heteropolysaccharides mainly composed of 4-ɑ-Glcp and also contain t-ɑ-Araf, 5-ɑ-Araf and 3,5-ɑ-Araf. They are spherical branched conformations without a triple-helix structure and can effectively increase the index of immune organs, lymphocyte proliferation, activate macrophages to regulate the immune system in mice and further enhance immune functions by improving delayed-type hypersensitivity reaction and antibody response. These results indicated that WGP-60-F and WGP-80-F could be used as potential immune enhancers, and gradient ethanol precipitation can be applied for the preparation of ginseng bioactive polysaccharide.

Effects of Extraction Methods on the Physicochemical Properties and Biological Activities of heteroglycans from Hibiscus sabdariffa calyx

The hot-water extraction method has been employed to enhance the diffusion rate of heteroglycans, thereby augmenting the extraction efficiency. This study investigates the antioxidant activities and immunological properties of water-soluble polysaccharides derived from the Hibiscus sabdariffa calyx (HWPF and LWPF). These polysaccharides were isolated through two distinct extraction methods involving hot water followed by ethanol precipitation. The physicochemical characterization was conducted through a series of experiments encompassing monosaccharide composition, ultraviolet-visible spectrum analysis, and Fourier-transform infrared (FT-IR) spectroscopy. The evaluation of antioxidative properties employed various techniques, including DPPH and ABTS radical scavenging activities, Ferric Reducing Antioxidant Power, ferrous chelating power, total reducing power, and protection against DNA damage. The assessment of anti-inflammatory potential involved the inhibition of hemolytic activity induced by a hypotonic solution and heat. Furthermore, the impact of the polysaccharides on the production of pro-inflammatory cytokines was analyzed through Luminex assay and RTPCR assay in PBMCs and LPS-stimulated RAW 264.7 cells, respectively. The findings revealed that heteroglycans (LWPF and HWPF) exhibit a content of Ara-Man-Rha-GluA-GalA (41.2:17.2:15.7:9.2:8.8) and Ara-Glu-Rha-Xyl-GluA-Man (31.5:23.8:12.16:10.4:7:6) with molecular weights of approximately 4.5 KDa and 41.3 KDa, respectively. HWPF demonstrated robust metal chelating and scavenging activities. Additionally, each fraction exhibited substantial protection of DNA against damage induced by H2O2. LWPF was observed to significantly down-regulate the expression of pro-inflammatory cytokines IL-6 and COX-2 at both transcriptional and translational levels, along with inhibiting NO production. In summary, these results indicate that the choice of extraction methods influences both the structure and biological activities of heteroglycans, providing valuable insights into the structure-activity relationship.
 Keywords: Hibiscus sabdariffa, heteroglycans, hot water extraction, physicochemical properties, Antioxidant activity, Pro-inflammatory cytokines.

Heteroglycan of Annona muricata leaves: Extraction, characterization, modulation of the inflammatory response in raw 264.7 macrophages and peripheral Blood mononuclear cells

The natural products contained in Annona muricata leaves are characterized by their ability to effectively enhance the immune response without exhibiting toxicity. The immunomodulatory effect of natural polysaccharides is considered one of the most important bioactive functions. In this study, we investigated the antioxidant activities and immunological properties of heteroglycans extracted from A. muricata leaves using hot water followed by ethanol precipitation. The polysaccharide was characterized for its physicochemical properties (UV spectroscopy, FTIR, gas chromatography), antioxidant potential (scavenging properties, chelating ability, reducing capability, and protection against DNA damage), immunomodulatory activities, and production of pro-inflammatory cytokines using peripheral blood mononuclear cells (PBMCs) and RAW 264.7 cells. The results indicated that the heteroglycans consisted of Glu-Man-GalA-GluA-Xyl and GluA-Man-Glu, respectively, for low and high molecular weight water-soluble polysaccharides coded as LWPC and HWPC. They exhibited strong metal chelating and scavenging activities. Additionally, each sample provided protection against DNA damage induction, with LWPC exhibiting up to 73.1 % protection. LWPC and HWPC significantly down-regulated the expression of pro-inflammatory cytokines TNF-α, IL-1β, IL-6, and COX-2, as well as NO production. At 200 µg/mL, only LWPC increased the production of IL-1β and IL-6 in PBMCs. Therefore, LWPC and HWPC hold promise for inhibiting radical-induced and inflammatory disorders.

Purification of exopolysaccharide produced from Lactobacillus spp. using ionic-liquid as adjuvant in alcohol/salt-based aqueous two-phase system for its antidiabetic properties

Diabetes is a chronic, metabolic disease characterized by hyperglycemia resulting from either insufficient insulin production or impaired cellular response to insulin. Exopolysaccharides (EPS) produced by Lactobacillus spp. demonstrated promising therapeutic potential in terms of their anti-diabetic properties. Extraction and purification of EPS produced by Lactobacillus acidophilus and Limosilactobacillus reuteri were performed using ethanol precipitation, followed by alcohol/salt based aqueous two-phase system (ATPS). The purification process involved ethanol precipitation followed by an alcohol/salt-based ATPS. The study systematically investigated various purification parameters in ATPS, including ethanol concentration, type and concentration of ionic liquid, type and concentration of salt and pH of salt. Purified EPS contents from L. acidophilus (63.30 μg/mL) and L. reuteri (146.48 μg/mL) were obtained under optimum conditions of ATPS which consisted of 30 % (w/w) ethanol, 25 % (w/w) dipotassium hydrogen phosphate at pH 10 and 2 % (w/w) 1-butyl-3-methylimidazolium octyl sulfate. The extracted EPS content was determined using phenol sulphuric acid method. In α-amylase inhibition tests, the inhibitory rate was found to be 92.52 % (L. reuteri) and 90.64 % (L. acidophilus), while in α-glucosidase inhibition tests, the inhibitory rate was 73.58 % (L. reuteri) and 68.77 % (L. acidophilus), based on the optimized parameters selected in ATPS. These results suggest that the purified EPS derived from the postbiotics of Lactobacillus spp. hold promise as potential antidiabetic agents.

Structure and immunostimulating activity of polysaccharides derived from the roots and leaves of dandelion

Two polysaccharides were obtained from dandelion roots (DPR) and dandelion leaves (DPL) via water extraction and ethanol precipitation. Both DPR and DPL were an acid heteropolysaccharide, with a molecular weight of 5.89 × 104 and 28.2 × 104 Da, respectively. Monosaccharide composition results showed that both DPR and DPL contained Man, Glc, Rha, GalA, Glc, Gal, and Ara with a molar ratios of 2.98:1.00:3.67:6.41:2.98:16.70:2.48 and 1.23:1.00:1.06:5.74:2.88:8.59:1.84, respectively. Methylation and NMR analysis showed DPR and DPL, with triple-helix conformations, were mainly composed of 4-ɑ-Galp and 4-ɑ-GalAp, terminated with t-ɑ-Araf, t-ɑ-Rhap, t-ɑ-Glcp and t-ɑ-Manp. DPL showed an immunoprotective effect in cyclophosphamide (Cy)-induced black-bone silky chickens by improving chicken growth performance, increasing the spleen, thymus, and bursa of Fabricius indices, and promoting blood lymphocyte proliferation, the secretion of cytokines (IL-2, IL-6, and INF-γ) and serum immunoglobulin (IgA, IgG, and IgM) levels in a dose-dependent manner. Moreover, the oxidative stress damage in immunosuppressed chickens was significantly reformed after DPL treatment. These findings provide useful information on the potential for application of dandelion polysaccharides as natural nutrients to enhance chicken immune and antioxidant functions.Graphical

A two-step method of cyclolinopeptide (linusorb) preparation from flaxseed cake via dry-screening

Linusorbs (LO), cyclolinopeptides, are a group of cyclic hydrophobic peptides and considered a valuable by-product of flaxseed oil due to numerous health benefits. Currently applied acetone or methanol extraction could contaminate the feedstocks for further food-grade application. Using flaxseed cake as feedstock, this study established a practical method for preparing LO from pressed cake. Firstly, LO composition of 15 flaxseed cultivars was analyzed. Next, cold-pressed cake was milled and screened mechanically. The kernel and hull fractions were separated based on the disparity of their mechanical strength. Monitored by hyperspectral fluorescence, the LO-enriched kernel fraction separated from cold-pressed flaxseed cake was further used as feedstock for LO production. After ethanol extraction, partition, and precipitation, LOs were extracted from cold-pressed flaxseed cake with a purity of 91.4%. The proposed method could serve as feasible flaxseed cake valorization strategy and enable the preparation of other polar compounds such as flax lignan and mucilage.

Hydrothermal processing of Sarcopeltis skottsbergii and study of the potential of its carrageenan for tissue engineering

The red macroalga Sarcopeltis skottsbergii was subjected to hydrothermal processing to maximize the solubilization and recovery of carrageenan. Once isolated by ethanol precipitation, the carrageenan was further chemically (oligosaccharides composition), and structurally (TGA/DTG, DSC, HPSEC, FTIR-ATR, 1H NMR, SEM, etc.) characterized, as well as employed as source for the synthesis of hydrogels. The rheological properties of the carrageenan showed promising results as biopolymer for food applications due to the high molecular weight (500 kDa) presenting higher cell viability than 70 %. The evaluation of immune activation using ELISA test reflected a lower inflammatory response for concentrations of 0.025 % of carrageenan. Conversely, the cell viability of the synthesized hydrogels did not surpass 50 %. This work represents a considerable step forward to obtain a biopolymer from natural sources and a thorough study of their chemical, structural and biological properties.

Comparison of immunomodulatory activity of polysaccharides and soluble dietary fibers and adsorption capacities of insoluble dietary fibers extracted from Lentinus edodes stipes

Stipes are the major waste from the processing of Lentinus edodes. To make full use of L. edodes stipes (LES), different fractions of LES polysaccharides (LESPs) were first obtained by water extraction and gradient ethanol precipitation. Afterwards, the LES residues were treated with an optimal combination of Aspergillus niger and Saccharomyces cerevisiae for the preparation of soluble/insoluble dietary fibers (LESS/LESI) using the response surface methodology and the Box-Behnken design. Subsequently, the in vitro immunomodulatory activity of LESPs and LESS, as well as the adsorption capacities of LESI were evaluated. The results showed that LESPs were neutral polysaccharides, mainly containing glucose. The optimal parameters for modifying the residues of LES were the followings: 4% (w/w) A. niger, 8% (w/w) S. cerevisiae, 31 ℃, 3 d, and a solid–liquid ratio of 1:12.5 in a yield of 14.73%/82.45% of LESS/LESI, respectively. The in vitro immunomodulatory activity assays revealed that LESPs and LESS had potent immunostimulatory activity to increase phagocytosis, acid phosphatase activity and nitric oxide production of RAW264.7 murine cell macrophages. The evaluation of adsorption capacities revealed that LESI owned stronger water holding capacity, oil holding capacity and water swelling capacity. This research could provide an effective way to fully utilize discarded L. edodes stipes with high added-value.Graphical An effective strategy was built to fully utilize discarded L. edodes stipes by preparing polysaccharides (LESPs-20, LESPs-50 and LESPs-80), soluble and insoluble dietary fibers (LESS/LESI). Their products showed immunomodulatory activity and adsorption capacities.

Protective effect of Ganoderma lucidum-fermented crop extracts against hydrogen peroxide- or β-amyloid-induced damage in human neuronal SH-SY5Y cells

BackgroundAlzheimer's disease (AD) is a neurodegenerative disorder characterized by the accumulation of stacked β-amyloid peptides in the brain and associated with the generation of oxidative stress. So far, there is no cure for AD or a way to stop its progression. Although the neuroprotective effects of Ganoderma lucidum aqueous extract and G. lucidum-derived triterpenoids and polysaccharides have been reported, the influence of G. lucidum-fermented crops on AD still lacks clarity.MethodsThis study aimed to investigate the protective effect of G. lucidum-fermented crop extracts against hydrogen peroxide- or β-amyloid peptide (Aβ25-35)-induced damage in human neuroblastoma SH-SY5Y cells.ResultsVarious extracts of G. lucidum-fermented crops, including extract A: 10% ethanol extraction using microwave, extract B: 70˚C water extraction, and extract C: 100˚C water extraction followed by ethanol precipitation, were prepared and analyzed. Extract B had the highest triterpenoid content. Extract C had the highest total glucan content, while extract A had the highest gamma-aminobutyric acid (GABA) content. The median inhibitory concentration (IC50, mg/g) for DPPH and ABTS scavenging activity of the fermented crop extracts was significantly lower than that of the unfermented extract. Pretreatment with these extracts significantly increased the cell viability of SH-SY5Y cells damaged by H2O2 or Aβ25-35, possibly by reducing cellular reactive oxygen species (ROS) and malondialdehyde (MDA) levels and increasing superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase (CAT) activities. Moreover, extract B markedly alleviated the activity of acetylcholinesterase (AChE), which is crucial in the pathogenesis of AD.ConclusionThese results clearly confirmed the effects of G. lucidum-fermented crop extracts on preventing against H2O2- or Aβ25-35-induced neuronal cell death and inhibiting AChE activity, revealing their potential in management of AD.

Characterization, antioxidant activity and in vitro digestion of hawthorn pectin prepared by gradient ethanol precipitation

Four modified hawthorn pectin fractions (MHPs), named MHP-30, MHP-50, MHP-70 and MHP-90, were obtained by ultrasonic-assisted pectin methyl esterase modification and gradient ethanol precipitation. The results indicated that all four MHPs were composed of galacturonic acid, galactose, xylose, arabinose, glucose and mannose in different proportions. With the increase of the ethanol concentration, the molecular weight, esterification degree and galacturonic acid content of MHPs all decreased, whereas the arabinose content and branching degree increased. The structural characterization from XRD, SEM, and FT-IR showed that four MHPs exhibited amorphous structure, similar functional groups, diverse surface morphologies. Besides, in vitro antioxidant assays confirmed that MHP-70 and MHP-90 exhibited stronger total antioxidant activities than MHP-30 and MHP-50. The results of simulated saliva-gastrointestinal digestion showed that the molecular weight of MHP-70 and MHP-90 remained stable, yielded small amounts of reducing sugars, and were resistant to digestion in the human upper digestive tract. Overall, MHP-70 and MHP-90 shown great potential as novel natural antioxidants, which are expected to be good carbon sources for the utilization of intestinal microorganisms.

Diversely regio-oxidative degradation of konjac glucomannan by lytic polysaccharide monooxygenase AA10 and generating antibacterial hydrolysate

Konjac glucomannan (KGM) hydrolysate exhibit various biological activities and health-promoting effects. Lytic polysaccharide monooxygenases (LPMOs) play an important role on enzymatic degradation of recalcitrant polysaccharides to obtain fermentable sugars. It is generally accepted that LPMOs exhibits high substrate specificity and oxidation regioselectivity. Here, a bacteria-derived SmAA10A, with chitin-active with strict C1 oxidation, was used to catalyse KGM degradation. Through ethanol precipitation, two hydrolysed KGM components (4 kDa (KGM-1) and 5 kDa (KGM-2)) were obtained that exhibited antibacterial activity against Staphylococcus aureus. In natural KGM, KGM-1, and KGM-2, the molar ratios of mannose to glucose were 1:2.19, 1:3.05, and 1:2.87, respectively, indicating that SmAA10A preferentially degrades mannose in KGM. Fourier-transform infrared spectroscopy and scanning electron microscopy imaging revealed the breakage of glycosylic bonds during enzymatic catalysis. The regioselectivity of SmAA10A for KGM degradation was determined based on the fragmentation behaviour of the KGM-1 and KGM-2 oligosaccharides and their NaBD4-reduced forms. SmAA10A exhibited diverse oxidation degradation of KGM and generated single C1-, single C4-, and C1/C4-double oxidised oligosaccharide forms. This study provides an alternative method for obtaining KGM degradation components with antibacterial functions and expands the substrate specificity and oxidation regioselectivity of bacterial LPMOs.

Immune-enhancing activity of compound polysaccharide on the inactivated influenza vaccine

Traditional Chinese medicine polysaccharides have numerous biological activities with broad applications in the biomedical industries. However, a clear understanding of the pharmacological activities of compound polysaccharides with multi-component structures remain challenging. This study aimed to investigate the immune boosting effect of compound polysaccharides on the influenza vaccine and assess the preliminary structure-activity relationship. The compound polysaccharide (CP) was isolated from the combined Chinese herbs lentinan, pachymaran and tremellan, and purified by gradient ethanol precipitation to obtain its subcomponents of CP-20, CP-40, CP-60, and CP-80 with decreasing molecular weights. These polysaccharides were mainly composed of glucans with different linkage patterns, including α-(1 → 3)-glucan, α-(1 → 4)-glucan and β-(1 → 6)-glucan. A significant improvement was observed in the survival of mice vaccinated with inactivated (IAV) vaccine and the isolated polysaccharides as adjuvants. A reduction in the pulmonary virus titer and weight loss were also observed. Moreover, CP-40 and CP-60, as well as the original CP, significantly enhanced the serum anti-IAV antibody titers and interleukin IL-2, IL-5, and IL-6 concentrations. These preliminary results indicate the immune boosting effect of the compound polysaccharides is highly relevant to the specific structural properties of the subcomponent, and CP-40 is worthy of further exploration as a glycan adjuvant for the IAV vaccine.

Effect of Hemicelluloses Fractionated by Graded Ethanol Precipitation from Corn Stover on the Enzymatic Hydrolysis of Lignocellulosic Biomass

Effect of Hemicelluloses Fractionated by Graded Ethanol Precipitation from Corn Stover on the Enzymatic Hydrolysis of Lignocellulosic Biomass

Unraveling umami complexity: From exploring umami peptides in fermented soybean curd to molecular elucidation of taste mechanisms

Sufu is a traditional umami fermented soybean curd (FSC). We isolated and identified 25 potential umami peptides using ethanol precipitation, dialysis desalination, resin adsorption recovery, nano-HPLC-MS/MS and virtual docking prediction. The results of sensory experiments showed that 23 of the synthetic peptides presented a lower umami threshold than that of MSG (0.50 mg/mL), and 10 peptides had obvious effects of increasing umami and saltiness. Seven peptides were quantified in FSC by UPLC-QQQ-MS, and the content of one novel umami tripeptide ADL (0.51 ± 0.03 mg/mL) was four times its umami threshold (0.125 mg/mL). Taste reconstitution experiments revealed that the umami peptides enhanced the umami and saltiness of the simulated system by 24.53% and 24.00%, respectively. Molecular dynamics proved that hydrogen bonding is the main interaction between peptide and receptors. This study is important for understanding and improving the taste quality of FSC.

Green synthesis of lichenan-decorated silver nanoparticles for catalytic hydrogenation of organic dyes and bacterial disinfection

Developing an eco-friendly and efficient homogeneous catalyst for treating complex wastewater containing toxic organic pollutants and bacteria is in significant demand. Herein, ultra-small Ag nanoparticles (NPs)-loaded random coil lichenan nanocomposite (L-AgNP) was prepared through a low-cost and green one-step in-situ reduction strategy using lichenan as the reducing agent. The random coil of lichenan acted as a micro-reactor to disperse and stabilize AgNPs via the Ag-O bonding interaction. In the participation of NaBH4, L-AgNP nanocomposite could achieve greater catalytic efficiency for catalyzing reduction of 4-nitrophenol (∼120 s, 34.34 × 10−3 s−1) with the smaller size of AgNPs or the higher content of Ag. The nanocatalyst also exhibited outstanding catalytic efficiency for congo red, rhodamine B, methylene blue, and methyl orange. The L-AgNP could be easily recycled through ethanol precipitation and continuously reused for ten cycles with a tiny loss of conversion rate. Furthermore, L-AgNP also displayed favorable antibacterial activity, inhibiting the growth of Escherichia coli and Staphylococcus aureus with more than 98 % disinfection ratio. This work provides a green strategy to develop polysaccharide-based multifunctional homogeneous catalysts focusing on wastewater purification.

Sequential Extraction and Characterization of Essential Oil, Flavonoids, and Pectin from Industrial Orange Waste.

Orange is one of the primary fruits processed into juice and other products worldwide, leading to a vast amount of waste accumulation. Such waste has been considered as an attractive candidate for upcycling to obtain bioactive components remaining. The present study investigated the extraction of essential oil (EO), flavonoids, and pectin from industrial orange waste with a holistic approach. To maximize EO yield and d-limonene concentration, hydrodistillation (HD) conditions were selected to be 5.5 mL water/g solid for 180 min. Remaining solids were further used for flavonoid extraction where conventional solvent, sequential ultrasound + solvent, and ultrasound-assisted extraction (UE) were applied. UE applied for 50 min with 120 mL solvent/g solid yielded the highest total phenolic (TPCs) and total flavonoid contents (TFCs), antioxidant capacity, and hesperidin and neohesperidin concentrations. In terms of TPC, TFC, antioxidant capacity, and antibacterial activity, both EO and flavonoid fractions demonstrated moderate to high bioactivity. At the final step, ethanol precipitation was applied to obtain the pectin that was solubilized in hot water during HD and it was characterized by Fourier transform infrared, degree of esterification, and galacturonic acid content. Practical application: to ensure utilization in the food, pharmaceutical, and cosmetic industries, this study presents a combined method to obtain several value-added compounds from industrial orange waste. Bioactive EO and flavonoids obtained could have applications in functional food, supplements, or cosmetic formulations, whereas extracted pectin can be used in many formulated foods and drugs.

Valorization of Nannochloropsis oceanica for integrated co-production of violaxanthin cycle carotenoids

The development of integrated co-production of multiple high-purity carotenoids from microalgal cells holds considerable significance for the valorization of microalgae. In this study, the economical microalga Nannochloropsis oceanica was identified as an accumulator of violaxanthin cycle carotenoids, including violaxanthin, antheraxanthin, and zeaxanthin. Notably, a novel and competent approach for the integrated co-production of violaxanthin cycle carotenoids was explored, encompassing four steps: microalgal cultivation, solvent extraction, octadecylsilyl open-column chromatography, and ethanol precipitation. Under optimal co-production conditions, the purities of the obtained violaxanthin, antheraxanthin, and zeaxanthin all exceeded 92%, with total recovery rates of approximately 51%, 40%, and 60%, respectively. Utilizing nuclear magnetic resonance techniques, the purified violaxanthin, antheraxanthin, and zeaxanthin were identified as all-trans-violaxanthin, all-trans-antheraxanthin, and all-trans-zeaxanthin, respectively. This method held significance for the multiproduct biorefinery of the microalga N. oceanica and carried potential future implications for the violaxanthin cycle carotenoids.

Structural Characterization and In Vitro Antioxidant Activity of a Novel Polysaccharide from Summer-Autumn Tea.

To enhance the utilization of summer-autumn tea, a water-soluble polysaccharide (D1N1) was isolated through a series of techniques including hot water extraction, ethanol precipitation, and column chromatography. The structure of D1N1 was determined through the utilization of ultraviolet, Fourier-transform infrared, high-performance anion-exchange chromatography, gas chromatography-tandem mass spectrometry, and nuclear magnetic resonance. The results revealed that glucose was the predominant component of D1N1, accounting for 95% of its composition. Additionally, D1N1 also contained galactose, arabinose, and rhamnose. The molecular weight (Mw) of D1N1 was determined to be 224.71 kDa. The backbone of D1N1 consisted of →4)-α-D -Glcp (1→, →3,4)-α-D-Galp-(1→, →4,6)-α-D -Glcp (1→ at a molar ratio of 35:1:1, and branching at the O-3 position of →3,4)-α-D-Galp-(1→ and O-6 position of →4,6)-α-D-Glcp (1→ with α-D -Glcp (1→. In addition, the antioxidant activity of D1N1 was also evaluated. D1N1 exhibited excellent antioxidant bioactivity against the DPPH, superoxide anion radical, and ABTS+ radical. These findings provide a theoretical basis for the application of summer-autumn tea polysaccharide as a potential functional food.

Novel Stable Protease Inhibitor from Phoenix dactylifera(L.) Flowers with Antimicrobial and Antitumoral Activities.

A novel protease inhibitor isolated from date palm Phoenix dactylifera(L.) flowers (PIDF) was purified and characterized. A heat and acidic treatment step followed by ethanol precipitation and reverse-phase high-performance chromatography was applied to purify this natural protease inhibitor to homogeneity with a single band of about 19 kDa. The stability study depicted that PIDF was fully stable at 40 °C and retained 65% of its initial activity after heating at 50 °C for 24 h. Its thermal stability at 70 °C was markedly enhanced by adding calcium, bovine serum albumin, and sorbitol as well as by metal divalent cations, especially Mg2+ and Hg2+. This protease inhibitor showed high inhibitory activity against therapeutic proteases, including pepsin, trypsin, chymotrypsin, and collagenase, and acted as a potent inhibitor of some commercial microbial proteases from Aspergillus oryzae, Bacillus. sp, and Bacillus licheniformis. Moreover, a potent antibacterial spectrum against Gram (+) and Gram (-) bacterial strains and an efficient antifungal effect were observed. Its cytotoxicity toward human colorectal cancer cell LoVo and HCT-116 lines suggested that PIDF could serve as a new therapeutic target inhibiting human colorectal cancer.