[Antioxidant activity of Euryale ferox seed shell extract and its therapeutic effects on oral ulcer in rats].

OBJECTIVE: To investigate the therapeutic effect of Euryale ferox seed shell extract on oral ulcer in rats and its underlying mechanism. METHODS: The contents of polyphenols and flavonoids in Euryale ferox seed shells were determined by Folin-phenol assay and aluminum nitrate colorimetry, respectively. DPPH·, ABTS+·, ·OH and·O2- scavenging experiments were performed to evaluate the antioxidant activities of Euryale ferox seed shell extract in vitro. In a rat model of oral ulcer induced by burning with glacial acetic acid, the therapeutic effect of Euryale ferox seed shell extract was assessed by detecting changes in serum levels of oxidative factors by enzyme-linked immunosorbent assay (ELISA) and observing pathological changes of the ulcerous mucosa using HE staining; the therapeutic mechanism of the extract was explored by detecting the expression levels of Keap1, Nrf2, Nes-Nrf2 and HO-1 proteins in ulcerous mucosa using Western blotting. RESULTS: The ethyl acetate extract of Euryale ferox seed shells contained 306.74±1.04 mg/g polyphenols and 23.43±0.61 mg/g flavonoids and had IC50 values for scavenging DPPH· and ABTS+· free radicals of 3.42 ± 0.97 µg/mL and 3.32 ± 0.90 µg/mL, respectively. In the rat models, the ethyl acetate extract significantly ameliorated oral mucosal ulcer, increased serum CAT level, and decreased serum MDA level. The protein expression levels of Nes-Nrf2 and HO-1 were increased and Keap1 protein expression was lowered significantly in the ulcerous mucosa of the rats after treatment with the extract (P<0.05 or 0.01). CONCLUSION: The therapeutic effect of Euryale ferox seed shell extract on oral ulcers in rats is mediated probably by activation of the Keap1/Nrf2/HO-1 signaling pathway.

The Ethnopharmacological, Phytochemical, and Pharmacological Review of Euryale ferox Salisb.: A Chinese Medicine Food Homology.

Euryale ferox Salisb. (prickly water lily) is the only extent of the genus Euryale that has been widely distributed in China, India, Korea, and Japan. The seeds of E. ferox (EFS) have been categorized as superior food for 2000 years in China, based on their abundant nutrients including polysaccharides, polyphenols, sesquineolignans, tocopherols, cyclic dipeptides, glucosylsterols, cerebrosides, and triterpenoids. These constituents exert multiple pharmacological effects, such as antioxidant, hypoglycemic, cardioprotective, antibacterial, anticancer, antidepression, and hepatoprotective properties. There are very few summarized reports on E. ferox, albeit with its high nutritional value and beneficial activities. Therefore, we collected the reported literature (since 1980), medical classics, database, and pharmacopeia of E. ferox, and summarized the botanical classification, traditional uses, phytochemicals, and pharmacological effects of E. ferox, which will provide new insights for further research and development of EFS-derived functional products.

Structure and physicochemical properties of low digestible Euryale ferox Salisb. seed starch.

BACKGROUND: Euryale ferox Salisb. is widely grown in China and Southeast Asia as a grain crop and medicinal plant. The composition, morphology, structure, physicochemical properties, thermal properties, and in vitro digestibility of North Euryale ferox seeds starch (NEFS), hybrid Euryale ferox seeds starch (HEFS), and South Euryale ferox seeds starch (SEFS) were studied. RESULT: Of the varieties that were studied, the amylose content of NEFS (23.03%) was the highest. Starch granules of each variety were smooth, sharp, small, and had an average diameter of 2 µm. All three varieties were A-type crystals with crystallinity ranging from 26.42% to 28.17%. The degree of double helix and the short-range order ranged from 1.9006 to 2.5324 and 1.4294 to 1.6006, respectively. The high proportion of C1 region in NEFS (17.74%) and HEFS (17.66%) were found. Thermodynamic properties in North Euryale ferox seeds included the highest onset temperature (To ) (71.43 °C), peak temperature (Tp ) (76.60 °C), conclusion temperature (Tc ) (82.77 °C), enthalpy of gelatinization (ΔH) (12.64 J g-1 ), and peak viscosity (1514 mPa·s). All three varieties maintained a low level of in vitro digestibility, with the highest resistant starch (RS) content (29.57%), the lowest rapidly digestible starch (RDS) content (27.07%), and the slowest hydrolysis kinetic constant (0.0303) in NEFS. CONCLUSION: The results revealed that the low digestibility of NEFS was attributable to compact granules, high crystallinity, high degree of order, and strong thermal stability. These digestive, physicochemical, and thermodynamic properties provide information for the future application of Euryale ferox seed starch in the food industry. © 2022 Society of Chemical Industry.

Valorization of unpopped Foxnut starch in stabilizing Pickering emulsion using OSA modification.

Starch was isolated from unpopped fox nut (Euryale ferox) and the effect of octenyl succinic anhydride (OSA) concentration (1, 2 and 3%) on physiochemical, functional, pasting, rheological and structural properties of was examined. The amylose content of native starch (22.9%) was higher than the modified starch (13.7%) for 3% OSA treatment. The water absorption capacity (1.29-0.9 g/g) significantly reduced, while oil absorption capacity (0.15-0.61 g/g), solubility (5-48%) and swelling power (2.77-13.60 g/g) increased after modification. The modification also altered the pasting properties by increasing the peak viscosity and reducing the pasting temperature. The cooked gel of all starch showed shear-thinning flow behavior and dynamic rheology confirmed reduction in storage and loss modulus after modification. Modified starch became rougher and irregular in shape and showed type A pattern as confirmed by SEM and XRD. Soybean oil-in-water Pickering emulsions were prepared by ultrasonication (US, 30 and 40% amplitude for 2 and 4 min) using starch as particle stabilizer and major factors influencing emulsion stability were investigated. Pickering emulsions prepared at 30 and 40% amplitude for 4 min US, produced the smaller droplet size, stable up to 15 days. However, all OSA modified starches were able to separate the oil and water even after the size of droplets increased with storage. The microstructure of the Pickering emulsions confirmed that starch particles aggregated in a tightly packed layer at the oil-water interface.

Study on structural characterization, physicochemical properties and digestive properties of euryale ferox resistant starch.

At present, the raw materials for industrialized RS3 products are relatively simple and its purity is low. In addition, the correlation between structure and digestion characteristics of RS3 are rarely studied. In this study, euryale ferox, a kind of annual aquatic herb crop with high content of starch was used as a raw material to prepare RS3 by different methods, including autoclaving, enzymolysis-autoclaving and dual enzymolysis, respectively. The results showed that there were significant differences in the structure and physicochemical properties of the different euryale ferox resistant-enhanced and purified resistant starches (p < 0.05). Purified euryale ferox resistant starches belonged to B + V type crystal and had high thermal stability. After digestion, the structure and thermal properties of euryale ferox resistant-enhanced starches changed a lot. The digestion rate and estimated glycemic index (eGI) of the three kinds of purified euryale ferox RS3 were lower than 20% and 50%, respectively.

Hepatoprotective Effect of Seed Coat ofEuryale ferox Extract in Non-alcoholic Fatty Liver Disease Induced by High-fat Diet in Mice by Increasing IRs-1 and Inhibiting CYP2E1.

Non-alcoholic fatty liver disease (NAFLD), a common chronic liver disease characterized by hepatic steatosis, affects 30-40% of the population in the world. The seed of Euryale ferox salisb. possesses several pharmacological actions, including metabolic syndrome. However, the seed coat of E. ferox was usually discarded as waste, which contains comparatively abundant polyphenols, and its biological activity has been rarely investigated. In this work, we evaluate the hepatoprotective effect of E. ferox seed coat extract (EFSCE), in NAFLD mice induced by high-fat diet (HFD). The HPLC-MS analysis indicated that the main components of EFSCE were polyphenols. And then, mice were treated with HFD for 4 weeks to induce NAFLD. The result showed that the body weight, weight of adipose tissue, the ratio of liver to body weight in NAFLD mice increased compared with control group. In addition, blood lipids parameters including total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL), low-density lipoprotein cholesterol (LDL) also increased in NAFLD mouse model. It was showed that, after treated with EFSCE (15 and 30 mg/kg/day) for 4 weeks, the body weight, lipids deposition in the liver and blood lipids in HFD-induced NAFLD mice markedly reduced. Compared with NAFLD mice, EFSCE administration could also prevent malondialdehyde (MDA) overproduction and strengthen Superoxide Dismutase (SOD) activity to counteract oxidative stress. Moreover, EFSCE was also found effective in reducing alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity in HFD-induced NAFLD model, which indicated liver injury in NAFLD. Therefore, EFSCE (rich in polyphenols) is indicated as bioactive nature product for HFD-induced NAFLD treatment, by eliminating lipid accumulation and oxidative stress via regulation of IRs-1 and CYP2E1.

Structural characterization and in vitro hypoglycemic activity of a glucan from Euryale ferox Salisb. seeds.

In this research, a polysaccharide fraction (EFSP-1) was obtained from the seeds of Euryale ferox Salisb. by DEAE sepharose FF and Superdex™ 75 gel chromatography. The average molecular weight (Mw) of EFSP-1 was 8.75 kDa. Monosaccharides composition analysis indicated that EFSP-1 was a glucan. The structure of EFSP-1 was characterized by analysis of FT-IR, GC-MS and NMR, which indicated that the backbone of EFSP-1 was mainly composed of (1→4)-α-D-Glcp with branches substituted at O-6 and terminated with T-α-D-Glcp. Moreover, the hypoglycemic effect of EFSP-1 was investigated by establishing insulin resistance HepG2 and 3T3-L1 cells. The results showed that EFSP-1 could increase glucose consumption by up-regulating the expression of GLUT-4 via activating PI3K/Akt signal pathway in IR cells. Hence, EFSP-1 could be a potential functional food to ameliorate insulin resistance for diabetes therapy.

Aquacultural, Nutritional and Therapeutic Biology of Delicious Seeds of Euryale ferox Salisb. : A Minireview.

BACKGROUND: Euryale ferox Salisb. (Makhana) is a rooted macro-hydrophyte, grown as a crop in north Bihar wetlands, India. Makhana refers to yummy pops prepared through mechanical smacking of the baked seeds of E. ferox. New techniques have been evolved to produce pops from seeds at a brisk pace. OBJECTIVE: It is specially associated with marital rituals possibly in view of its spermatogenic properties. It has a high amino acid index but a low glycemic value. The plant carries medicinal values against cardiovascular and diabetes-related diseases. Makhana pops with a high volume to mass ratio, is developed as a reliable system for drug delivery in the recent times. RESULTS: There is a fair prospect of raising multi-faceted industry based on Makhana culture, harvest, post-harvest safety, preparation of pops and various types of food products therefrom and marketing in areas where water-logging is considered as a curse. It may generate employment and strengthen regional economy propping check on population migration primarily due to non-employment and poverty. CONCLUSION: Active research is required to harness this macro-hydrophyte along with co-culture of compatible fish varieties and other hydrophytes into a sustainable livelihood and economic prosperity.

Relationship between the flavonoid composition and flower colour variation in Victoria.

Victoria (Nymphaeaceae), an annual or perennial aquatic plant genus, contains only two species: V. amazonica (Poepp.) J. C. Sowerby and V. cruziana A. D. Orb. Both species have large floating leaves and variable flower colour. Both Victoria species are night bloomers, which have white petals on the first blooming night that then turn pink or ruby red on the second blooming day. The mechanism of the colour change of Victoria petals during anthesis is still unclear. In this study, flavonoids in Victoria petals of both species were evaluated and quantified by high-performance liquid chromatography with photodiode array detection (HPLC-DAD) and by ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS) for the first time. In total, 14 flavonoids were detected in Victoria petals, including 4 anthocyanins and 10 flavonols. The flavonoid compositions differed across the two species, resulting in different colours between the inner and outer petals. With increased anthocyanin content across blooming days, the colour of Victoria flowers changed over time. The results of this study will improve understanding of the chemical mechanism of colour formation and lay the foundation for selective colour breeding in Victoria.

Extract of Euryale ferox Salisb exerts antidepressant effects and regulates autophagy through the adenosine monophosphate-activated protein kinase-UNC-51-like kinase 1 pathway.

Positive regulation of autophagy by Euryale ferox Salisb (ES) leads to antidepressant effects. This study focused on the potential antidepressant mechanisms induced by the petroleum ether fraction of ES (ES-PE) in the chronically unpredictable mild stress (CUMS) mouse model. Deficits in the sucrose preference test, open field test, tail suspension test, and forced swim test were observed following CUMS, and were reversed following ES-PE administration. Contrary to the expression of mammalian target of rapamycin, autophagy was decreased after establishment of the CUMS model. We also observed trends for downregulation of adenosine mononphosphate-activated protein kinase (AMPK) and mammalian autophagy-initiating kinase (ULK1), which were differentially affected by ES-PE. HT22 cells and Compound C, an inhibitor of AMPK, were used to verify the results in mice. Gas chromatography-mass spectrometry analysis revealed high level of vitamin E acetate in ES-PE. Taken together, our data indicate that ES-PE activated autophagy by regulating the AMPK pathway. © 2018 IUBMB Life, 70(4):300-309, 2018.

Prophylactic effects of alkaloids from Ba lotus seeds on L-NNA-induced hypertension in mice.

Alkaloids from Ba lotus seeds (ABLS) are a kind of important functional compounds in lotus seeds. The present study was designed to determine its hypertension prophylactic effects in the L-NNA-induced mouse hypertension model. The mice were treated with ABLS, the serum and tissues levels of NO, MDA, ET-1, VEGF, and CGRP were determined using the experimental kits, the mRNA levels of various genes in the heart muscle and blood vessel tissues were further determined by RT-PCR assay. ABLS could reduce the systolic blood pressure (SBP), mean blood pressure (MBP), and diastolic blood pressure (DBP), compared to that of the model control group. After ABLS treatment, the NO (nitric oxide) contents in serum, heart, liver, kidney and stomach of the mice were higher than that of the control mice, but the MDA (malonaldehyde) contents were lower than that of the control mice. The serum levels of ET-1 (endothelin-1), VEGF (vascular endothelial growth factor) were decreased after ABLS treatment, but CGRP (calcium gene related peptide) level was increased. The ABLS treated mice had higher mRNA expressions of HO-1, nNOS, and eNOS and lower expressions of ADM, RAMP2, IL-1ß, TNF-α, and iNOS than the control mice. Higher concentration of ABLS had greater prophylactic effects, which were close to that of the hypertension drug captopril. These results indicated the hypertension prophylactic effects of ABLS could be further explored as novel medicine or functional food in the future.

[Analysis and Evaluation of Inorganic Elements in Euryale Semen from Different Habitats by Microwave Digestion-ICP-OES].

OBJECTIVE: To establish a simple, rapid and efficient method for determination of different inorganic elements in Euryale Semen from different habitats. METHODS: Inductively coupled plasma-optical emission spectrometry(ICP-OES) was applied to determine inorganic elements in Euryale Semen, and the results were analyzed by principal component analysis. RESULTS: Euryale Semen from different habitats contained the kind of inorganic elements ranging from 22 to 26, including micronutrient elements like Iron, Zinc, Selenium, Copper, Molybdenum, Chrome and Cobalt, as well as macronutrient elements such as Potassium, Calcium, Sodium, Magnesium and Phosphorus. Five factors were extracted and used to comprehensively evaluate Euryale Semen from 20 different habitats covered almost China. The comprehensive function was F = 0. 38828F1 + 0. 25603F2 + 0. 07617F3 + 0. 06860F4 + 0. 04868F5, which resulted in the top three samples coming from Jiangsu Gaoyou, Hunan Xiangxi and Jiangsu Suzhou respectively. CONCLUSION: The study indicates that ICP-OES is a quick, accurate and sensitive method to determine the contents of inorganic elements in Euryale Semen,which provides scientific and reliable reference for its quality control and safety assessment.

Simultaneous Determination of 16 Nucleosides and Nucleobases in Euryale ferox Salisb. by Liquid Chromatography Coupled with Electro Spray Ionization Tandem Triple Quadrupole Mass Spectrometry (HPLC-ESI-TQ-MS/MS) in Multiple Reaction Monitoring (MRM) Mode.

In this study, a simple, rapid, efficient analytical method was established for the qualification and quantification of 16 nucleosides and nucleobases in Euryale ferox Salisb. by using liquid chromatography coupled with electrospray ionization tandem triple quadrupole mass spectrometry (HPLC-ESI-TQ-MS/MS) in multiple-reaction monitoring (MRM) mode. Ideal separation of 16 target compounds was achieved on Xbridge Amide HILIC column (4.6 × 150 mm, 3.5 µm) with gradient elution in 11 min by optimized conditions. Variations of nucleosides and nucleobase in samples from different cultivation regions ranging from 190.50 to 1594.30 µg/g were obvious. The total nucleoside contents were higher than total nucleobases, especially in the compositions of guanosine, cytidine and 2'-deoxyguanosine. Samples 1-18 with dense thorns were better characters than samples 19-26 without thorns in terms of nucleosides and nucleobases concentrations in general. The limits of detection (LODs) and quantification (LOQs) for 16 analytical substances were investigated to be 0.11-6.33 ng/mL and 0.35-21.1 ng/mL, respectively. And the method was first applied to large aquatic plants with good linearity, precision, repeatability and accuracy. All present information provided a scientific and rational reference for quality assessment and control of Euryale ferox Salisb.

[Studies on chemical constituents from seeds of Euryale ferox].

OBJECTIVE: To study the chemical constituents from the seeds of Euryale ferox. METHODS: The chemical constituents were isolated by silica gel column, Sephadex LH-20 and their structures were identified by physico-chemical and spectral analysis. RESULTS: Seven compounds were purified from the 95% ethanol extract. These constituents were elucidated as protocatechuic acid (1), gallic acid (2), gallic acid ethyl ester(3),5 ,7-dihydroxychromone(4), ß-sitosterol(5), daucosterol(6), and 5,7-dihydroxy-6,4'-dimethoxyflavone(7), respectively. CONCLUSION: All compounds are isolated from this plant for the first time.

Antioxidant and anti-fatigue activities of phenolic extract from the seed coat of Euryale ferox Salisb. and identification of three phenolic compounds by LC-ESI-MS/MS.

This study investigated the antioxidant potential and anti-fatigue effects of phenolics extracted from the seed coat of Euryale ferox Salisb. The in vitro antioxidant potentials, including scavenging DPPH, hydroxyl radical activities and reducing power were evaluated. Antioxidant status in vivo was analyzed by SOD, CAT, GSH-Px activities and the MDA content in liver and kidneys of D-galactose-induced aging mice. The anti-fatigue effect was evaluated using an exhaustive swimming test, along with the determination of LDH, BUN and HG content. The phenolic extract possessed notable antioxidant effects on DPPH, hydroxyl radical scavenging and reducing power. The mice which received the phenolic extract showed significant increases of SOD, CAT (except for in the kidney), GSH-Px activities, and a decrease of MDA content. The average exhaustive swimming time was obviously prolonged. Meanwhile, increase of LDH content and decrease of BUN content were observed after mice had been swimming for 15 min. The HG storage of mice was improved in the high and middle dose extract groups compared with the normal group. The contents of total phenols and gallic acid of the extract were determined. Three compounds in the extract were identified as 5,7-dihydroxy-2-(3,4,5-trihydroxyphenyl)-chroman-4-one, 5,7,4-trihydroxyflavanone and buddlenol E. These results suggest that the extract of E. ferox is a promising source of natural antioxidants and anti-fatigue material for use in functional foods and medicines.

Two new tocopherol polymers from the seeds of Euryale ferox.

Two new tocopherol polymers, chroman-type dimer named ferotocodimer A (1) and spiro-type trimer named ferotocotrimer E (2), were isolated from the seeds of Euryale ferox. Their structures were elucidated on the basis of spectroscopic methods including HR-ESI-MS and 1D and 2D NMR. The absolute configurations of 1 and 2 were determined by CD and ROESY experiments.

Isolation and identification of compounds responsible for antioxidant capacity of Euryale ferox seeds.

Euryale ferox seed is consumed medicinally or for food in China. The present study revealed it to contain significant antioxidant activity, which may be associated with its medical applications as a proteinuria inhibitor of diabetic nephropathy. This study resulted in the identification of 3 new sesquineolignans, named euryalins A-C (1-3), and 16 known compounds, which were all first isolated from this plant apart from 5,7,4-trihydroxy-flavanone. The antioxidant potential of the partial isolates was evaluated using the DPPH radical scavenging assay and mesangial cellular assay. Compounds 2, rel-(2α,3ß)-7-O-methylcedrusin (4), syringylglycerol-8-O-4-(sinapyl alcohol) ether (5), and (+)-syringaresinol (7) were found to be most active on DPPH assay, whereas compounds 2, 4, 7, (1R,2R,5R,6S)-2-(3,4-dimethoxyphenyl)-6-(3,4-dihydroxyphenyl)-3,7-dioxabicyclo[3.3.0]octane, and buddlenol E could significantly inhibit high glucose-stimulated reactive oxygen species production in mesangial cells. The results suggested that E. ferox seed could be considered as an excellent source of natural antioxidants and is useful in the prevention of diabetic nephropathy.

Cerebrosides and tocopherol trimers from the seeds of Euryale ferox.

Two new cerebrosides, ferocerebrosides A (1) [(2S,3R,4E,8E,2'R)-1-O-(beta-glucopyranosyl)-N-(2'-hydroxydocosanoyl)-4,8-sphingadienine] and B (2) [(2S,3R,4E,8E,2'R)-1-O-(beta-glucopyranosyl)-N-(2'-hydroxytetracosanoyl)-4,8-sphingadienine], two new tocopherol trimers, ferotocotrimers C (5) and D (6), and two known tocopherol trimers, IVb (3) and IVa (4), were isolated from the seeds of Euryale ferox. Their structures were determined on the basis of spectroscopic data, especially 1D and 2D NMR experiments. Compounds 1 and 2 showed cytotoxicity in the brine shrimp lethality bioassay, with LC50 values of 0.17 and 0.20 mM, respectively.

The effect of Euryale ferox (Makhana), an herb of aquatic origin, on myocardial ischemic reperfusion injury.

Fox nut or gorgon nut (Euryale ferox--Family Nymphaeaceae), popularly known as Makhana, has been widely used in traditional oriental medicine to cure a variety of diseases including kidney problems, chronic diarrhea, excessive leucorrhea and hypofunction of the spleen. Based on the recent studies revealing antioxidant activities of Euryale ferox and its glucosides composition, we sought to determine if Euryale ferox seeds (Makhana) could reduce myocardial ischemic reperfusion injury. Two different models were used: acute model, where isolated rat hearts were preperfused for 15 min with Krebs Henseleit bicarbonate (KHB) buffer containing three different doses of makhana (25, 125 or 250 microg/ml) followed by 30 min of ischemia and 2 h of reperfusion; and chronic model, where rats were given two different doses of makhana (250 and 500 mg/kg/day) for 21 days, after which isolated hearts were subjected to 30 min of ischemia followed by 2 h of reperfusion. In both cases, the hearts of the Makhana treated rats were resistant to ischemic reperfusion injury as evidenced by their improved post-ischemic ventricular function and reduced myocardial infarct size. Antibody array technique was used to identify the cardioprotective proteins. The Makhana-treated hearts had increased amounts of thioredoxin-1 (Trx-1) and thioredoxin-related protein-32 (TRP32) compared to the control hearts. Western blot analysis confirmed increased expression of TRP32 and thioredoxin proteins. In vitro studies revealed that Makhana extracts had potent reactive oxygen species scavenging activities. Taken together, the results of this study demonstrate cardioprotective properties of Makhana and suggest that such cardioprotective properties may be linked with the ability of makhana to induce TRP32 and Trx-1 proteins and to scavenge ROS.

[The effects of pronuciferine on NO and NOS production in cultured human umbilical vein endothelium cells].

OBJECTIVE: To investigate the effects of Pronuciferine on nitric oxide (NO), total nitric oxide synthase (tNOS) and inducible nitric oxide synthase (iNOS) production in cultured Human Umbilical Vein Endothelium Cells (HUVECs). METHODS: Pronuciferine was extracted from plumula nelumbinis. The experiments were performed in HUVECs in vitro. Cell-morphology was observed by light microscope. Cells viability was assessed by MTT assay. NO, tNOS and iNOS were measured by Colorimetry. RESULTS: Cell- morphology and viability weren't affected by Pronuciferine. Pronuciferine significantly increased the level of NO and the activity of tNOS, but had no effect on the activity of iNOS. CONCLUSION: Pronuciferine increases the level of NO by the enhancement of the activity of tNOS. It may have the protection on endothelial function.