Showing papers by "Tuba Esatbeyoglu published in 2022"
TL;DR: In this paper , the authors used UPLC-MS to assess metabolites heterogeneity among four major Cinnamomum species, including true cinnamon and less explored species (C. tamala).
Abstract: The present study aimed to assess metabolites heterogeneity among four major Cinnamomum species, including true cinnamon (Cinnamomum verum) and less explored species (C. cassia, C. iners, and C. tamala). UPLC-MS led to the annotation of 74 secondary metabolites belonging to different classes, including phenolic acids, tannins, flavonoids, and lignans. A new proanthocyanidin was identified for the first time in C. tamala, along with several glycosylated flavonoid and dicarboxylic fatty acids reported for the first time in cinnamon. Multivariate data analyses revealed, for cinnamates, an abundance in C. verum versus procyandins, dihydro-coumaroylglycosides, and coumarin in C. cassia. A total of 51 primary metabolites were detected using GC-MS analysis encompassing different classes, viz. sugars, fatty acids, and sugar alcohols, with true cinnamon from Malaysia suggested as a good sugar source for diabetic patients. Glycerol in C. tamala, erythritol in C. iners, and glucose and fructose in C. verum from Malaysia were major metabolites contributing to the discrimination among species.
16 citations
TL;DR: In this article , the authors investigated the cytotoxicity of basil at three different growth stages (GS), i.e., GS-1, GS-2, and GS-3 using the brine shrimp lethality assay.
Abstract: Basil (Ocimum basilicum L.) is one of the most common aromatic herbs, a rich source of bioactive compounds, and is used extensively to add aroma and flavor to food. The leaves, both in fresh and dried form, are used as a culinary ingredient in different cultures. O. basilicum is also famous for its therapeutic potential and preservation effects. The present study investigated the cytotoxicity of basil at three different growth stages (GS), i.e., GS-1 (58 days of growth), GS-2 (69 days of growth), and GS-3 (93 days of growth) using the brine shrimp lethality assay. The results revealed that cytotoxicity was influenced by GS and the concentration of extracts. Aqueous extracts of basil at a concentration of 10 to 1000 µg/mL did not show notable toxicity. The lowest mortality rate, i.e., 8.9%, was recorded for GS-2 at the highest tested dose of basil extracts. The mortality rate at GS-1, GS-2, and GS-3 was found to be 26.7 ± 3.34%, 8.91 ± 0.10%, and 16.7 ± 0.34%, respectively, at 1000 µg/mL. GS-2 basil powder with the lowest toxicological risk was extracted with different solvents, viz., n-hexane, dichloromethane, ethanol, and water. The highest concentration of plant secondary metabolites including total phenolic acid, flavonoids, and tannin content was observed in ethanol extracts. Ethanol extracts also exhibited the highest antioxidant activity in DPPH, FRAP and H2O2 assays. LC-ESI-MS/MS analysis presented ethanol extracts of basil as a promising source of known health-promoting and therapeutic compounds such as rosmarinic acid, ellagic acid, catechin, liquiritigenin, and umbelliferone. The results suggest basil, a culinary ingredient, as a potential source of bioactive compounds which may offer an array of health promoting and therapeutic properties.
15 citations
TL;DR: In this article , the authors explored the biodiversity of lactic acid microflora and characterization of LAB is a new approach to form a variety of starter communities to create innovative nutritional food matrices.
Abstract: Lactic acid bacteria (LAB) are common microorganisms found in various ecosystems including in plants, fermented foods, and the human body. Exploring the biodiversity of lactic acid microflora and characterization of LAB is a new approach to form a variety of starter communities to create innovative nutritional food matrices. There has been growing interest in LAB isolated from non-dairy environments as these bacteria exhibit significant metabolic diversity and have unique taste-forming activities. Disease may be prevented, or treated by LAB but the treatment of disease conditions with LAB is highly dependent on the host's microbiome and diet and varies in both effectiveness and side effects from individual to individual. Future perspectives on the study of LAB may be related to the expansion of our knowledge in the fields of genetics and genetic engineering. The application of genetic science may help to improve existing strains and develop new strains with characteristics designed for specific purposes. Therefore, the preservative effects of LAB and their metabolites, as well as their interaction on the growth of food borne pathogens and food spoilage microorganisms were elucidated. In addition, the competitive models for microbial growth between LAB and other microorganisms as well as the role of LAB in the elimination of toxic compounds in food products were discussed. Moreover, the review provided an overview of the risks and benefits of using LAB in the food industry.
10 citations
TL;DR: It can be stated that T. terrestris is a potential bearing herb and findings of current study further verify the claims made in ancient medicinal systems, however, after investigation of each identified compound, it must be considered for drug discovery.
Abstract: Tribulus terrestris L. belongs to the family Zygophyllaceae and integral part of various ancient medicinal systems including Chinese, Indian, and European to combat various health ailments. The aim of the present study was to assess the phytochemical constituents, in vitro antioxidant activity using DPPH, FRAP, and H2O2 assays, in vitro anticancer activity using MTT assay, and in vitro and in vivo anti-inflammatory properties of T. terrestris. The acute and sub-acute toxicity of extracts exhibiting most biological potential was examined using murine models. Liquid–liquid partitioning followed by RP–HPLC sub-fraction of crude extract was performed. After that, ESI-MS/MS analysis was done for the timid identification of bioactive metabolites responsible for bioactivities of sub-fractions and HPLC analysis to quantify the compounds using external standards. Among all extracts, T. terrestris methanol extract was noted to hold maximum phenolic (341.3 mg GAE/g) and flavonoid (209 mg QE/g) contents, antioxidant activity in DPPH (IC50 71.4 µg/mL), FRAP (35.3 mmol/g), and H2O2 (65.3% inhibition) assays, anti-inflammatory activities in vitro at 400 µg/mL (heat-induced hemolysis, % inhibition 68.5; egg albumin denaturation, % inhibition 75.6%; serum albumin denaturation, % inhibition 80.2), and in vivo at 200 mg/kg (carrageenan-induced paw edema, % inhibition 69.3%; formaldehyde-induced paw edema, % inhibition 71.3%) and anticancer activity against breast cancer cell (MCF-7) proliferation (IC50 74.1 µg/mL). Acute and sub-acute toxicity studies recorded with no change in body weight, behavior, hematological, serum, and histopathological parameters in treated rats with T. terrestris methanol extracts when compared to control group. Fraction B obtained through liquid–liquid partitioning resulted in more bioactive potential as compared to the parent methanol extract. RP–HPLC analysis of fraction B resulted with four sub-fractions (TBTMF1-TBTMF4), wherein TBTMF3 delineated notable bioactive capabilities as compared to other fractions and parent methanol extract. ESI-MS/MS analysis of TBTMF3 resulted with tentative identification of myricetin, rutin, liquitrigenin, physcion, and protodioscin. It can be stated that T. terrestris is a potential bearing herb and findings of current study further verify the claims made in ancient medicinal systems. However, after investigation of each identified compound, it must be considered for drug discovery.
9 citations
TL;DR: In this paper , the most effective extract, only consisting of anthocyanins, copigments or a mixture of both, obtained from a lingonberry juice concentrate was identified.
Abstract: Lingonberries contain high contents of bioactive compounds such as chlorogenic acids and anthocyanins. In addition to radical scavenging and antioxidant activities, these compounds can protect cells from DNA damage. For this reason, lingonberries might be well suited for nutraceuticals or natural biomedicines. To assess these applications, the present study characterized and identified the most effective extract, only consisting of anthocyanins, copigments or a mixture of both, obtained from a lingonberry juice concentrate. An extract was generated by using a XAD-7 column followed by fractionation into anthocyanins and copigments using adsorptive membrane chromatography. After identification of main polyphenols by HPLC–photodiode array–electrospray ionization–tandem mass spectrometry, free radical scavenging activity was analyzed by electron spin resonance spectroscopy using 2,2-diphenyl-1-picrylhydrazyl and galvinoxyl radicals. Furthermore, cyclic voltammetry analyses and the Trolox equivalent antioxidant capacity (TEAC) assay were applied. Finally, the reactive oxygen species (ROS) reducing effects of the lingonberry extract and its fractions were evaluated in HepG2 cells. While the combination of anthocyanins and copigments possessed the highest antioxidant activities, all samples (XAD-7 extract, anthocyanin and copigment fraction) protected cells from oxidative stress. Thus, synergistic effects between phenolic compounds may be responsible for the high antioxidant potential of lingonberries, enabling their use as nutraceuticals.
9 citations
TL;DR: In this paper , a concise explanation of quercetin's action mechanism and potential health applications is given, along with a detailed analysis of the potential applications of the compound in terms of anti-cancer properties.
Abstract: Naturally occurring phytochemicals with promising biological properties are quercetin and its derivatives. Quercetin has been thoroughly studied for its antidiabetic, antibacterial, anti-inflammatory, anti-Alzheimer's, anti-arthritic, antioxidant, cardiovascular, and wound-healing properties. Anticancer activity of quercetin against cancer cell lines has also recently been revealed. The majority of the Western diet contains quercetin and its derivatives, therefore consuming them as part of a meal or as a food supplement may be sufficient for people to take advantage of their preventive effects. Bioavailability-based drug-delivery systems of quercetin have been heavily studied. Fruits, seeds, vegetables, bracken fern, coffee, tea, and other plants all contain quercetin, as do natural colors. One naturally occurring antioxidant is quercetin, whose anticancer effects have been discussed in detail. It has several properties that could make it an effective anti-cancer agent. Numerous researches have shown that quercetin plays a substantial part in the suppression of cancer cells in the breast, colon, prostate, ovary, endometrial, and lung tumors. The current study includes a concise explanation of quercetin's action mechanism and potential health applications.
8 citations
TL;DR: In this paper , the authors summarize the most recent literature related to botany, traditional applications, phytochemical ingredients, pharmacological activities, nutrition, and potential food applications of Syzygium cumini.
Abstract: Syzygium cumini, locally known as Jamun in Asia, is a fruit-bearing crop belonging to the Myrtaceae family. This study aims to summarize the most recent literature related to botany, traditional applications, phytochemical ingredients, pharmacological activities, nutrition, and potential food applications of S. cumini. Traditionally, S. cumini has been utilized to combat diabetes and dysentery, and it is given to females with a history of abortions. Anatomical parts of S. cumini exhibit therapeutic potentials including antioxidant, anti-inflammatory, analgesic, antipyretic, antimalarial, anticancer, and antidiabetic activities attributed to the presence of various primary and secondary metabolites such as carbohydrates, proteins, amino acids, alkaloids, flavonoids (i.e., quercetin, myricetin, kaempferol), phenolic acids (gallic acid, caffeic acid, ellagic acid) and anthocyanins (delphinidin-3,5-O-diglucoside, petunidin-3,5-O-diglucoside, malvidin-3,5-O-diglucoside). Different fruit parts of S. cumini have been employed to enhance the nutritional and overall quality of jams, jellies, wines, and fermented products. Today, S. cumini is also used in edible films. So, we believe that S. cumini’s anatomical parts, extracts, and isolated compounds can be used in the food industry with applications in food packaging and as food additives. Future research should focus on the isolation and purification of compounds from S. cumini to treat various disorders. More importantly, clinical trials are required to develop low-cost medications with a low therapeutic index.
8 citations
TL;DR: In this article , the effects of non-thermal treatments and food matrix on the bioaccessibility and transepithelial transportation of phenolics from cranberrybush were investigated, and the results showed that PEF15 treatment enhanced the recovery of total flavonoids (TFC; increase of 3.9% ± 1.1), chlorogenic acid (increase of 29.9%) and antioxidant capacity after gastrointestinal digestion.
8 citations
TL;DR: In this article , the authors evaluated the effect of single and coupled processing methods on the chemical composition, Antioxidant Activity and Lipid Quality of Tiger Nuts (Cyperus esculentus L).
Abstract: The objective of this study was to evaluate the effect of single and coupled processing methods on the chemical composition, Antioxidant Activity and Lipid Quality of Tiger Nuts (Cyperus esculentus L). The processing methods applied were oven and sun drying, boiling water, steaming, blanching and roasting. After processing, tiger nut powders were analyzed for their total phenolic content, antioxidant activity, oil quality, nutrients and anti-nutrients composition. Results showed that boiling and blanching significantly (p<0.05) reduced the total phenolic content of tiger nuts (24.66 to 20.17 mg GAE/100 g for boiling and 39.23 to 30.26 GAE/100 g for blanching) while this parameter increased with roasting (40.73 to 300.44 mg GAE/100 g). Drying coupled to roasting significantly (p<0.05) increased the antioxidant activity (from 11.00-17.40 to 56.34-96.33%) of the nuts. The analysis of oils showed that the treatments applied significantly (p<0.05) increased its peroxide (PV) (from 1.69 to 21.31 meq O2/Kg) and thiobarbituric acid (TBA) (0.32 to 3.77 ppm) values. Oil from roasting for 10 min was the most altered with PV and TBA values of 21.31 meq O2/Kg and 2.23 ppm, respectively. The analysis of the proximate composition showed that the processing methods used have significantly (p<0.05) reduced the carbohydrate (from 52.13 to 36.57%) and protein (from 11.73 to 4.58%) contents of tiger nuts while the lipid content increased (from 26.49 to 42.56%). Processed Tiger nuts were found to be rich in phosphorus, sodium, potassium, calcium and magnesium. All the processing methods have significantly reduced (p<0.05) the anti-nutrient content of tiger nuts.
7 citations
TL;DR: The role of flaxseed substances in the maintenance of a healthy composition of the gut microbiome, prevention, and management of multiple diseases has recently been elucidated in various studies, which have highlighted its importance further as a powerful nutritional remedy as discussed by the authors .
Abstract: Flaxseed (Linum usitatissimum L.) has gained worldwide recognition as a health food because of its abundance in diverse nutrients and bioactive compounds such as oil, fatty acids, proteins, peptides, fiber, lignans, carbohydrates, mucilage, and micronutrients. These constituents attribute a multitude of beneficial properties to flaxseed that makes its use possible in various applications, such as nutraceuticals, food products, cosmetics, and biomaterials. The importance of these flaxseed components has also increased in modern times because of the newer trend among consumers of greater reliance on a plant-based diet for fulfilling their nutritional requirements, which is perceived to be hypoallergenic, more environmentally friendly, sustainable, and humane. The role of flaxseed substances in the maintenance of a healthy composition of the gut microbiome, prevention, and management of multiple diseases has recently been elucidated in various studies, which have highlighted its importance further as a powerful nutritional remedy. Many articles previously reported the nutritive and health benefits of flaxseed, but no review paper has been published reporting the use of individual flaxseed components in a manner to improve the techno-functional properties of foods. This review summarizes almost all possible applications of flaxseed ingredients in food products from an extensive online literature survey; moreover, it also outlines the way forward to make this utilization even better.
7 citations
TL;DR: In this article , the empagliflozin (EMPA) hepatoprotective efficacy in thioacetamide (TAA)-induced hepatotoxicity by targeting AMPK/SIRT-1 activity and reducing HIF-1α was discovered.
Abstract: Hepatic fibrosis causes severe morbidity and death. No viable treatment can repair fibrosis and protect the liver until now. We intended to discover the empagliflozin’s (EMPA) hepatoprotective efficacy in thioacetamide (TAA)-induced hepatotoxicity by targeting AMPK/SIRT-1 activity and reducing HIF-1α. Rats were treated orally with EMPA (3 or 6 mg/kg) with TAA (100 mg/kg, IP) thrice weekly for 6 weeks. EMPA in both doses retracted the serum GGT, ALT, AST, ammonia, triglycerides, total cholesterol, and increased serum albumin. At the same time, EMPA (3 or 6 mg/kg) replenished the hepatic content of GSH, ATP, AMP, AMPK, or SIRT-1 and mitigated the hepatic content of MDA, TNF-α, IL-6, NF-κB, or HIF-1α in a dose-dependent manner. Likewise, hepatic photomicrograph stained with hematoxylin and eosin or Masson trichrome stain of EMPA (3 or 6 mg/kg) revealed marked regression of the hepatotoxic effect of TAA with minimal injury. Similarly, in rats given EMPA (3 or 6 mg/kg), the immunohistochemically of hepatic photomicrograph revealed minimal stain of either α-SMA or caspase-3 compared to the TAA group. Therefore, we concluded that EMPA possessed an antifibrotic effect by targeting AMPK/SIRT-1 activity and inhibiting HIF-1α. The present study provided new insight into a novel treatment of liver fibrosis.
TL;DR: In this paper , the essential oil composition, antioxidant activity and in vitro cytotoxic capacity of the Ocimum basilicum essential oil (OB-EO) of basil indigenous to Pakistan was investigated, and the results suggest OB-EO as a promising carrier of bioactive compounds with a broad range of food preservation properties.
Abstract: A commonly observed chicken meat issue is its lipid oxidation that leads to deterioration of its organoleptic and nutritional properties and its further-processed products. Basil (Ocimum basilicum L.) is one of the traditional culinary herbs exhibiting food preservation properties. The current study investigated the essential oil composition, antioxidant activity and in vitro cytotoxic capacity of the essential oil of basil indigenous to Pakistan. GC–MS analysis of the essential oil revealed the presence of 59 compounds that constituted 98.6% of the essential oil. O. basilicum essential oil (OB-EO) exhibited excellent antioxidant activity, i.e., IC50 5.92 ± 0.15 µg/mL as assayed by the DPPH assay, 23.4 ± 0.02 µmoL Fe/g by FRAP, and 14.6 ± 0.59% inhibition by H2O2. The brine shrimp lethality assay identified an average mortality of ~18% with OB-EO at 10–1000 µg/mL, while that of the same concentration range of the standard drug (etoposide) was 72%. OB-EO was found to be non-toxic to HeLa and PC-3 cell lines. TBARS contents were significantly decreased with increase of OB-EO in chicken nuggets. The lowest TBARS contents were recorded in nuggets supplemented with 0.3% OB-EO, whereas the highest overall acceptability score was marked to the treatments carrying 0.2% OB-EO. The results suggest OB-EO as a promising carrier of bioactive compounds with a broad range of food preservation properties, and which has a sensory acceptability threshold level for chicken nuggets falling between 0.2-0.3% supplementation. Future research must investigate the antibacterial impact of OB-EO on meat products preserved with natural rather than synthetic preservatives.
TL;DR: The promising ability of SG-loaded nanosystem to ameliorate insulin resistance, protect against oxidative stress, and restore pancreatic β-cell secretory function warrants its inclusion in further studies during diabetes progression.
Abstract: β-Sitosterol glucoside (SG), isolated from Senecio petasitis (Family Asteraceae), was loaded in self-nanoemulsifying drug delivery systems (SEDDS) in a trial to enhance its solubility and biological effect. Various co-surfactants were tested to prepare a successful SEDDS. The selected SG-loaded SEDDS had a droplet size of 134 ± 15.2 nm with a homogenous distribution (polydispersity index 0.296 ± 0.02). It also demonstrated a significant augmentation of SG in vitro release by 4-fold compared to the free drug suspension. The in vivo insulin sensitivity and antidiabetic effect of the prepared SG-loaded SEDDS were further assessed in streptozotocin-induced hyperglycemic rats. The hypoglycemic effect of SG-loaded nanosystem was evidenced by decreased serum glucose and insulin by 63.22% and 53.11%, respectively. Homeostasis model assessment-insulin resistance (HOMA-IR) index demonstrated a significant reduction by 5.4-fold in the diabetic group treated by SG-loaded nanosystem and exhibited reduced glucagon level by 40.85%. In addition, treatment with SG-loaded nanosystem significantly decreased serum MDA (malondialdehyde) and increased catalase levels by 38.31% and 64.45%, respectively. Histopathological investigations also supported the protective effect of SG-loaded nanosystem on the pancreas. The promising ability of SG-loaded nanosystem to ameliorate insulin resistance, protect against oxidative stress, and restore pancreatic β-cell secretory function warrants its inclusion in further studies during diabetes progression.
TL;DR: In this paper , the total phenolic content, anthocyanins, phenolic acids, antioxidant capacity and α-amylase inhibitory activity of black (Aydin Siyahi), purple (Kadife Kemer), and white (Trabzon Kadife) eggplants grown in Turkey were subjected to a comparative investigation.
Abstract: The total phenolic content, anthocyanins, phenolic acids, antioxidant capacity and α-amylase inhibitory activity of black (Aydin Siyahi), purple (Kadife Kemer) and white (Trabzon Kadife) eggplants grown in Turkey were subjected to a comparative investigation. The black cultivar exhibited the highest total phenolic (17,193 and 6552 mg gallic acid equivalent/kg fw), flavonoid (3019 and 1160 quercetin equivalent/kg fw) and anthocyanin (1686 and 6167 g delphinidin-3-O-glucoside equivalent/kg fw) contents in crude extracts of the peel and pulp. The majority of the caffeic acid was identified in the ester (2830 mg/kg fw) and ester-bound (2594 mg/kg fw) forms in the peel of ‘Kadife Kemer’ and in the glycoside form (611.9 mg/kg fw) in ‘Aydin Siyahi’, as well as in the pulp of these two eggplants. ‘Kadife Kemer’ (purple eggplant) contained the majority of the chlorogenic acid in free form (27.55 mg/kg fw), compared to ‘Aydin Siyahi’ in the ester (7.82 mg/kg fw), glycoside (294.1 mg/kg dw) and ester-bound (2.41 mg/kg fw) forms. The eggplant cultivars (peel and pulp, mg/kg fw) exhibited a relatively high delphinidin-3-O-rutinoside concentration in the peel of ‘Aydin Siyahi’ (avg. 1162), followed by ‘Kadife Kemer’ (avg. 336.6), and ‘Trabzon Kadife’ (avg. 215.1). The crude phenolic extracts of the eggplants exhibited the highest antioxidant capacity values (peel and pulp, µmoL Trolox equivalent/kg fw) of 2,2-diphenyl-1-picrylhydrazyl (DPPH, 8156 and 2335) and oxygen radical absorbance capacity (ORAC, 37,887 and 17,648). The overall results indicate that black and purple eggplants are the cultivars with greater potential benefits in terms of their phenolics and antioxidant values than the white eggplant.
TL;DR: In this paper , the authors reviewed the nutritional and phytochemical profile as well as biological activities exhibited by the apple seed and highlighted the current application of apple seed in food, pharmaceutical and nutraceutical industries along with the safety aspects considering the presence of amygdalin.
Abstract: Recent trends in food industry focus on utilization of fruit processing waste and by-products as novel ingredients due to their nutritional benefits. Apple seed is one such novel ingredient that can find application as functional ingredient in food, nutraceutical and pharmaceutical industry. The existing studies suggest that apple seed is a rich source of amino acids, fatty acids and polyphenolic compounds. This review discusses the nutritional and phytochemical profile as well as biological activities exhibited by the apple seed. It also highlights the current application of apple seed in food, pharmaceutical and nutraceutical industries along with the safety aspects considering the presence of amygdalin. The apple seed meal, extracts or seed oil as novel ingredient can find various industrial applications given their dense nutritional profile. Besides their functionality, rigorous and extensive research is required to promote its usage at industrial level for harnessing the benefits of apple seeds. • This is pioneering work on reviewing phytochemical profile and biological activities of apple seed. • Apple seed is a superior source of bioactive compounds. • Apple seed extracts display cardioprotective, antidiabetic, & anticancer activities. • Antioxidant and antimicrobial activities make apple seed suitable for use in foods.
TL;DR: In this paper , the effects of using natural lignocellulosic-based adsorbents from sugarcane bagasse (SC), cornstalk piths (CP), and corn cob (CC) on the physicochemical properties and quality of fried oils were studied.
Abstract: In this work, the effects of using natural lignocellulosic-based adsorbents from sugarcane bagasse (SC), cornstalk piths (CP), and corn cob (CC) on the physicochemical properties and quality of fried oils were studied. The properties of lignocellulosic biomasses were examined using X-ray diffraction (XRD), scanning electron microscope (SEM), and Fourier transform infrared spectroscopy (FTIR). Moreover, the changes in the physicochemical properties of fresh, fried oils (for 4, 8, 12, 16 and 20 h) and adsorbents-treated oils were examined. The XRD results revealed that SC and CP biomasses have more amorphous regions than CC biomass, which had the highest crystallinity percentage. The results also showed that lignocellulosic biomasses enhanced the quality of the used oils. SC was the most effective biomass to enhance the properties of the used sunflower oil. For instance, the acid value of oil samples fried for 20 h reduced from 0.63 ± 0.02 to 0.51 ± 0.02 mg KOH/g oil after SC biomass treatment. For the peroxide value, the SC biomass treatment reduced it from 9.45 ± 0.56 (fried oil for 20 h) to 6.91 ± 0.12 meq O2/kg. Similarly, SC biomass adsorbent reduced the p-Anisidine Value (p-AV) of the used oil (20 h) from 98.45 ± 6.31 to 77.92 ± 3.65. Moreover, SC adsorbents slightly improved the lightness of the used oils (20 h). In conclusion, natural lignocellulosic biomasses, particularly SC, could be utilized as natural adsorbents to improve the oil quality. The results obtained from this study could help in developing sustainable methods to regenerate used oils using natural and cheap adsorbents.
TL;DR: In this article , the effects of the Supercritical Anti-Solvent (SAS) process and food models on the antioxidant capacity, bioaccessibility and transport dynamics of PVP based microparticles were investigated using a combined in vitro gastrointestinal digestion/Caco-2 cell culture model.
Abstract: In the present study, the effects of the Supercritical Anti-Solvent (SAS) process and food models on the antioxidant capacity, bioaccessibility and transport dynamics of flavonol-loaded polyvinylpyrrolidone (PVP) based microparticles were investigated using a combined in vitro gastrointestinal digestion/Caco-2 cell culture model. SAS-processed and unprocessed flavonols were supplied in two different food models: 10% ethanol for an aqueous hydrophilic food simulant and 3% acetic acid for an acidic food simulant. The SAS processing of quercetin and rutin resulted in a much higher recovery of these bioactives as well as greater retention of antioxidant capacity after gastrointestinal digestion in both hydrophilic and acidic food models. The present study also demonstrates that SAS coprecipitation has a positive effect on the stability and transport of bioactives across the epithelial cell layer. It can be deduced from the results that the SAS process can be a useful method in pharmaceutical and nutraceutical applications with high stability, bioaccessibility, bioavailability and thus enhanced nutritional value.
TL;DR: It is proposed that CLEE may reduce TAA-hepatorenal toxicity via its antioxidant and anti-inflammatory properties suppressing oxidative stress and alleviation of plasma liver enzymes, kidney function parameters, and inflammatory markers.
Abstract: Cyperus species represent a group of cosmopolitan plants used in folk medicine to treat several diseases. In the current study, the phytochemical profile of Cyperus laevigatus ethanolic extract (CLEE) was assessed using UPLC-QTOF–MS/MS. The protective effect of CLEE at 50 and 100 mg /kg body weight (b.w.) was evaluated on hepatorenal injuries induced by thioacetamide (100 mg/kg) via investigation of the extract’s effects on oxidative stress, inflammatory markers and histopathological changes in the liver and kidney. UPLC-QTOF–MS/MS analysis of CLEE resulted in the identification of 94 compounds, including organic and phenolic acids, flavones, aurones, and fatty acids. CLEE improved the antioxidant status in the liver and kidney, as manifested by enhancement of reduced glutathione (GSH) and coenzyme Q10 (CoQ10), in addition to the reduction in malondialdehyde (MDA), nitric oxide (NO), and 8-hydroxy-2′-deoxyguanosine (8OHdG). Moreover, CLEE positively affected oxidative stress parameters in plasma and thwarted the depletion of hepatorenal ATP content by thioacetamide (TAA). Furthermore, treatment of rats with CLEE alleviated the significant increase in plasma liver enzymes, kidney function parameters, and inflammatory markers. The protective effect of CLEE was confirmed by a histopathological study of the liver and kidney. Our results proposed that CLEE may reduce TAA-hepatorenal toxicity via its antioxidant and anti-inflammatory properties suppressing oxidative stress.
TL;DR: In this article , a review on the application of Traceability 4.0 in the fruits and vegetables sector, focusing on relevant Industry 4-0 enablers, especially Artificial Intelligence, the Internet of Things, blockchain, and big data, is presented.
TL;DR: In this paper, a study was conducted to improve nutritional, functional and consumer safety aspects of cabbage powder (CP) by using microwave heating, blanching, alkali or acid washing treatments.
Abstract: This study was aimed to improve nutritional, functional and consumer safety aspects of cabbage powder (CP). White cabbage (Brassica oleracea var. capitata f. alba) was dehydrated to CP following microwave heating, blanching, alkali or acid washing treatments. The results for nutrients and mineral composition of raw and processed CP elucidated raw CP to exhibit significantly (p < 0.05) higher amounts of protein (12.2%), dietary fiber (25.2%), Na (52 mg/100 g), Ca (355 mg/100 g), K (286 mg/100 g), Fe (14 mg/100 g) and Zn (32 mg/100 g). Among different processing techniques, microwave treatment resulted in a higher rate of reduction for alkaloids, oxalates, tannins and phytates contents, i.e., 77%, 85%, 85%, and 86%, respectively. Likewise, microwave treatment was found more effective in reducing residual levels of neonicotinoids, pyrethroids, organophosphates including imidacloprid, cypermethrin, bifenthrin, chlorpyrifos and deltamethrin in cabbage in the range of 0.98–0.12 ppm, 1.22–0.23 ppm, 1.03–0.15 ppm, 1.97–0.43 ppm, and 2.12–0.36 ppm, respectively. CP supplementation at the rate of 5% in unleavened flatbreads was observed to maintain textural and sensory attributes of the product. The results suggest microwave heating as a cost-effective technique to reduce toxicants load in cabbage powder. Further, ~5% supplementation of CP in wheat flour may also improve nutritional and functional properties of the baked goods.
TL;DR: In this paper , the authors investigated the phytochemical composition, biological activities, and in silico molecular docking of the ethanolic extract of Zaleya pentandra L. jaffery.
Abstract: Zaleya pentandra L. jaffery is the only species of the genus Zaleya that grows in the Cholistan desert, Pakistan. It is a Xero-halophyte plant with high phenolic and flavonoid content. The present research was designed to investigate the phytochemical composition, biological activities, and in silico molecular docking of the ethanolic extract of Z. pentandra. The phytochemical evaluation was done through preliminary phytochemical testing, estimation of total bioactive content, and gas chromatography–mass spectrometry (GC–MS) analysis for the identification of volatile compounds. For the evaluation of biological activities, antioxidants, and enzyme inhibition (α-glucosidase, cholinesterase, and tyrosinase), antibacterial and antiviral assays were performed. GC–MS analysis revealed the presence of 29 tentative volatile compounds. The ethanolic extract of Z. pentandra contains high phenolic content (119.6 ± 0.12 mg GAE/g extract) and flavonoid content (45.5 ± 0.19 mg QE/g extract), which correlates with the strong DPPH, FRAP, and enzyme inhibition results. The ethanolic extract of Z. pentandra also showed dose-dependent antibacterial activity. Micrococcus luteus and Pseudomonas aeruginosa were found to be most susceptible, with 16 mm and 17 mm zone of inhibitions at a maximum dose of 20 mg/mL. Antiviral results showed that the ethanol extract has excellent activity against H9, IBV, and NDV viral strains. Additionally, in silico molecular docking was performed in order to determine the interaction and binding affinity between the enzymes and compounds identified by GC–MS. α-glucosidase, cholinesterase, and tyrosinase showed the highest binding affinity toward 1,2-benzenedicarboxylic acid, 2-hydroxy-n-(2-phenylethyl) benzamide, γ-sitosterol, and lactose. These findings can serve as a benchmark for anti-diabetic-, neuro-, and skin-protective uses of this plant and can be used for the isolation of pure bioactive compounds in the future.
TL;DR: In this paper , a GC-MS analysis of tea tree oil (TTO) revealed 38 volatile components with sesquiterpene hydrocarbons (43.56%) and alcohols (41.03%) as major detected classes.
Abstract: The GC-MS analysis of tea tree oil (TTO) revealed 38 volatile components with sesquiterpene hydrocarbons (43.56%) and alcohols (41.03%) as major detected classes. TTO efficacy is masked by its hydrophobicity; nanoencapsulation can address this drawback. The results showed that TTO-loaded solid lipid nanoparticles (SLN1), composed of glyceryl monostearate (2% w/w) and Poloxamer188 (5% w/w), was spherical in shape with a core-shell microstructure. TTO-SLN1 showed a high entrapment efficiency (96.26 ± 2.3%), small particle size (235.0 ± 20.4 nm), low polydispersity index (0.31 ± 0.01), and high negative Zeta potential (−32 mV). Moreover, it exhibited a faster active agent release (almost complete within 4 h) compared to other formulated TTO-SLNs as well as the plain oil. TTO-SLN1 was then incorporated into cellulose nanofibers gel, isolated from sugarcane bagasse, to form the ‘TTO-loaded nanolipogel’ which had a shear-thinning behavior. Second-degree thermal injuries were induced in Wistar rats, then the burned skin areas were treated daily for 7 days with the TTO-loaded nanolipogel compared to the unmedicated nanolipogel, the TTO-loaded conventional gel, and the normal saline (control). The measurement of burn contraction proved that TTO-loaded nanolipogel exhibited a significantly accelerated skin healing, this was confirmed by histopathological examination as well as quantitative assessment of inflammatory infiltrate. This study highlighted the success of the proposed nanotechnology approach in improving the efficacy of TTO used for the repair of skin damage induced by burns.
TL;DR: In this paper , the in vitro bioavailability of rosehip infusion phenolics, mainly catechin, as a response to conventional and non-thermal treatments by combining gastrointestinal digestion and a Caco-2 cell culture model, was investigated.
Abstract: In this study, the in vitro bioavailability of rosehip infusion phenolics, mainly catechin, as a response to conventional and non-thermal treatments by combining gastrointestinal digestion and a Caco-2 cell culture model, was investigated. After application of thermal treatment (TT, 85 °C/10 min), high pressure (HPP, 600 MPa/5 min) or pulsed electric field (PEF, 15 kJ/kg) processing, all samples were subjected to simulated gastrointestinal digestion. Then, the amount of maximum non-toxic digest ratio was determined by the cytotoxicity sulforhodamine B (SRB) assay. Next, Caco-2 cells were exposed to 1:5 (v/v) times diluted digests in order to simulate the transepithelial transportation of catechin. Results showed that non-thermally processed samples (5.19 and 4.62% for HPP and PEF, respectively) exhibited greater transportation across the epithelial cell layer compared to than that of the TT-treated sample (3.42%). The present study highlighted that HPP and PEF, as non-thermal treatments at optimized conditions for infusions or beverages, can be utilized in order to enhance the nutritional quality of the final products.
TL;DR: D. juxtapostia root methanol extract showed improvement in hepatic cell structure and low cellular infiltration but, in contrast, the dichloromethane extract did not show any significant improvement in hepatocyte morphology, cellular infiltration, or necrosis of hepatocytes in comparison to the positive control.
Abstract: An estimated 450 species of Dryopteris in the Dryoperidaceae family grow in Japan, North and South Korea, China, Pakistan, and Kashmir. This genus has been reported to have biological capabilities; however, research has been conducted on Dryopteris juxtapostia. Therefore, with the present study, we aimed to exploring the biological potential of D. juxtapostia root and shoot extracts. We extracted dichloromethane and methanol separately from the roots and shoots of D. juxtapostia. Antioxidant activity was determined using DPPH, FRAP, and H2O2 assays, and anti-inflammatory activities were evaluated using both in vitro (antiurease activity) and in vivo (carrageenan- and formaldehyde-induced paw edema) studies. Toxicity was evaluated by adopting a brine shrimp lethality assay followed by determination of cytotoxic activity using an MTT assay. Hepatoprotective effects of active crude extracts were examined in rats. Activity-bearing compounds were tentatively identified using LC-ESI-MS/MS analysis. Results suggested that D. juxtapostia root dichloromethane extract exhibited better antioxidant (DPPH, IC50 of 42.0 µg/mL; FRAP, 46.2 mmol/g; H2O2, 71% inhibition), anti-inflammatory (urease inhibition, 56.7% at 50 µg/mL; carrageenan-induced edema inhibition, 61.7% at 200 µg/mL; formaldehyde-induced edema inhibition, 67.3% at 200 µg/mL), brine shrimp % mortality (100% at 1000 µg/mL), and cytotoxic (HeLa cancer, IC50 of 17.1 µg/mL; prostate cancer (PC3), IC50 of 45.2 µg/mL) effects than D. juxtapostia root methanol extract. D. juxtapostia shoot dichloromethane and methanol extracts exhibited non-influential activity in all biological assays and were not selected for hepatoprotective study. D. juxtapostia root methanol extract showed improvement in hepatic cell structure and low cellular infiltration but, in contrast the dichloromethane extract, did not show any significant improvement in hepatocyte morphology, cellular infiltration, or necrosis of hepatocytes in comparison to the positive control, i.e., paracetamol. LC-ESI-MS/MS analysis showed the presence of albaspidin PP, 3-methylbutyryl-phloroglucinol, flavaspidic acid AB and BB, filixic acid ABA and ABB, tris-desaspidin BBB, tris-paraaspidin BBB, tetra-flavaspidic BBBB, tetra-albaspidin BBBB, and kaempferol-3-O-glucoside in the dichloromethane extract, whereas kaempferol, catechin, epicatechin, quinic acid, liquitrigenin, and quercetin 7-O-galactoside in were detected in the methanol extract, along with all the compounds detected in the dichloromethane extract. Hence, D. juxtapostia is safe, alongside other species of this genus, although detailed safety assessment of each isolated compound is obligatory during drug discovery.
TL;DR: In this paper , the occurrence of Staphylococcus aureus in chicken fillets and to control its growth using various lyophilized seaweed extracts (i.e., Halimeda opuntia (HO), Actinotrichia fragilis, and Turbinaria turbinata) was assessed by an agar disk diffusion assay in vitro.
Abstract: This study aims to assess the occurrence of Staphylococcus aureus in chicken fillets and to control its growth using various lyophilized seaweed extracts (i.e., Halimeda opuntia (HO), Actinotrichia fragilis, and Turbinaria turbinata) by an agar disk diffusion assay in vitro. Results showed that prevalence of S. aureus in breast and thigh samples reached of 92% and 84%, respectively. Lyophilized HO extract was the only seaweed that showed the antibacterial activity against S aureus with a significant difference at p < 0.05. The minimum inhibitory concentration (MIC) of HO extract was 1.5%, with an inhibition zone of 8.16 ± 0.73 mm. Regarding 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity, IC50 was recorded at 55.36 μg/mL, whereas cytotoxic IC50 of the lyophilized HO extract on peripheral blood mononuclear cells (PBMCs) was 33.7 µg/mL; a higher IC50 of HO extracts permits their use as a safe food additive in meat products. Moreover, total phenolic compounds and total flavonoids compounds recorded 20.36 ± 0.092 and 16.59 ± 0.029 mg/mL, respectively. HPLC analyses of phenolic compounds profiles exhibited many bioactive substances and the higher ratio was daidzein with 10.84 ± 0.005 µg/mL and followed by gallic acid with a value of 4.06 ± 0.006 µg/mL. In a challenge study, chicken fillet (CHF) experimentally inoculated with S. aureus (ST) and treated with the lyophilized HO algal extract at 4% and 6% (CHF/ST/HO) showed a complete reduction of S. aureus count on the 6th and 4th days in chicken fillet stored at 4 °C, respectively. Moreover, CHF/ST/HO at 4% and 6% of HO extract enhanced the sensory attributes of grilled un-inoculated chicken fillet. Thus, lyophilized HO extracts are promising antibacterial and antioxidant candidates in the chicken meat industry.
TL;DR: Averrhoa carambola L. is reported for its anti-obese and anti-diabetic activities as discussed by the authors , which improved all obesity complications and exhibited significant activity in an obese rat model.
Abstract: Averrhoa carambola L. is reported for its anti-obese and anti-diabetic activities. The present study aimed to investigate its aqueous methanol leaf extract (CLL) in vivo anti-obese activity along with the isolation and identification of bioactive compounds and their in vitro α-glucosidase inhibition assessment. CLL improved all obesity complications and exhibited significant activity in an obese rat model. Fourteen compounds, including four flavone glycosides (1–4) and ten dihydrochalcone glycosides (5–12), were isolated and identified using spectroscopic techniques. New compounds identified in planta included (1) apigenin 6-C-(2-deoxy-β-D-galactopyranoside)-7-O-β-D-quinovopyranoside, (8) phloretin 3′-C-(2-O-(E)-cinnamoyl-3-O-β-D-fucopyranosyl-4-O-acetyl)-β-D-fucopyranosyl-6′-O-β-D fucopyranosyl-(1/2)-α-L arabinofuranoside, (11a) phloretin3′-C-(2-O-(E)-p-coumaroyl-3-O-β-D-fucosyl-4-O-acetyl)-β-D-fucosyl-6′-O-(2-O-β-D-fucosyl)-α-L-arabinofuranoside, (11b) phloretin3′-C-(2-O-(Z)-p-coumaroyl-3-O-β-D-fucosyl-4-O-acetyl)-β-D-fucosyl-6′-O-(2-O-β-D-fucosyl)-α-L-arabinofuranoside. Carambolaside M (5), carambolaside Ia (6), carambolaside J (7), carambolaside I (9), carambolaside P (10a), carambolaside O (10b), and carambolaside Q (12), which are reported for the first time from A. carambola L. leaves, whereas luteolin 6-C-α-L-rhamnopyranosyl-(1-2)-β-D-fucopyranoside (2), apigenin 6-C-β-D-galactopyranoside (3), and apigenin 6-C-α-L-rhamnopyranosyl-(1-2)-β-L-fucopyranoside (4) are isolated for the first time from Family. Oxalidaceae. In vitro α-glucosidase inhibitory activity revealed the potential efficacy of flavone glycosides, viz., 1, 2, 3, and 4 as antidiabetic agents. In contrast, dihydrochalcone glycosides (5–11) showed weak activity, except for compound 12, which showed relatively strong activity.
TL;DR: In this article , a stable active microemulgel containing flaxseed extract was developed and incorporated in Carbopol-940 gel matrix to formulate an active micro emulsion with 59.15% release in in vitro studies.
Abstract: Antioxidants from natural sources are extensively attaining consideration to avert the skin from damage and aging caused by free radicals. Flaxseed (Linum usitatissimum L.), a natural therapeutic agent, was meant to be explored cosmeceutical by quantifying its potential phytoconstituents and to be incorporated into a microemulgel for topical use. Hydroalcoholic fractions (both methanolic and ethanolic; 80%) flaxseed extracts were subjected to phytochemical screening by quantifying total phenolic content (TPC), total flavonoid content (TFC), and high-performance liquid chromatography-ultraviolet (HPLC-UV), and for biological activities through 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, tyrosinase inhibition assay, and sun protection factor (SPF). Ethanolic fraction was selected for further study by TPC (18.75 mg gallic acid equivalent/g) and TFC (1.34 mg quercetin equivalent/g). HPLC-UV analysis showed the existence of benzoic, quercetin, caffeic, vanillic, p-coumaric, gallic, cinnamic, syringic, and sinapic acids. Biological activities showed 87.00%, 72.00%, and 21.75 values for DPPH assay, tyrosinase inhibition, and SPF assays, respectively. An oil-in-water (OW) microemulsion containing the flaxseed extract, with 99.20 nm Zeta size, −19.3 Zeta potential and 0.434 polydispersity index was developed and incorporated in Carbopol-940 gel matrix to formulate an active microemulgel with 59.15% release in in vitro studies. The successfully formulated stable active microemulgel produced statistically significant effects (p < 0.05), in comparison to a placebo, on skin erythema, melanin, sebum, moisture, and elasticity, in a noninvasive in vivo study performed on 13 healthy human female volunteers. Other cosmeceutical products can also be formulated from flaxseed, making it a considerable candidate for further utilization in the pharmaceutical industry.
TL;DR: In this paper , the degradation efficiency of natural organic acid extracts (i.e., citric, malic, tartaric, and lactic) against mycobacterial complex containing mycolic acids (MAs) was evaluated in vitro in phosphate buffer solution (PBS) and Karish cheese models.
Abstract: Degradation of the mycobacterial complex containing mycolic acids (MAs) by natural bioactive compounds is essential for producing safe and value-added foods with therapeutic activities. This study aimed to determine the degradation efficiency of natural organic acid extracts (i.e., citric, malic, tartaric, and lactic), quadri-mix extract from fruits and probiotics (i.e., lemon, apple, grape, and cell-free supernatant of Lactobacillus acidophilus), and synthetic pure organic acids (i.e., citric, malic, tartaric, and lactic), against MA in vitro in phosphate buffer solution (PBS) and Karish cheese models. The degradation effect was evaluated both individually and in combinations at different concentrations of degradants (1, 1.5, and 2%) and at various time intervals (0, 6, 12, 24, and 48 h). The results show that MA degradation percentage recorded its highest value at 2% of mixed fruit extract quadri-mix with L. acidophilus and reached 99.2% after 48 h both in PBS and Karish cheese, unlike other treatments (i.e., citric + malic + tartaric + lactic), individual acids, and sole extracts at all concentrations. Conversely, organic acid quadri-mix revealed the greatest MA degradation% of 95.9, 96.8, and 97.3% at 1, 1.5, and 2%, respectively, after 48 h. Citric acid was more effective in MA degradation than other acids. The fruit extract quadri-mix combined with L. acidophilus-fortified Karish cheese showed the highest sensorial characteristics; hence, it can be considered a novel food-grade degradant for MA and could be a promising biocontrol candidate against Mycobacterium tuberculosis (Mtb) in food matrices.
TL;DR: In this article , the adsorption abilities of activated charcoal (A. charcoal), yeast (Saccharomyces cerevisiae), and the probiotic Lactobacillus rhamnosus as AFs adsorbents in three forms (sole, di-and tri-mix) were evaluated.
Abstract: The high incidence of aflatoxins (AFs) in chocolates suggests the necessity to create a practical and cost-effective processing strategy for eliminating mycotoxins. The present study aimed to assess the adsorption abilities of activated charcoal (A. charcoal), yeast (Saccharomyces cerevisiae), and the probiotic Lactobacillus rhamnosus as AFs adsorbents in three forms—sole, di- and tri-mix—in phosphate-buffered saline (PBS) through an in vitro approach, simulated to mimic the conditions present in the gastrointestinal tract (GIT) based on pH, time and AFs concentration. In addition, the novel fortification of chocolate with A. charcoal, probiotic, and yeast (tri-mix adsorbents) was evaluated for its effects on the sensory properties. Using HPLC, 60 samples of dark, milk, bitter, couverture, powder, and wafer chocolates were examined for the presence of AFs. Results showed that all the examined samples contained AFs, with maximum concentrations of 2.32, 1.81, and 1.66 µg/kg for powder, milk, and dark chocolates, respectively. The combined treatment demonstrated the highest adsorption efficiency (96.8%) among all tested compounds. Scanning electron microscope (SEM) analysis revealed the tested adsorbents to be effective AF-binding agents. Moreover, the novel combination of tri-mix fortified chocolate had a minor cytotoxicity impact on the adsorptive abilities, with the highest binding at pH 6.8 for 4 h, in addition to inducing an insignificant effect on the sensory attributes of dark chocolate. Tri-mix is thus recommended in the manufacturing of dark chocolate in order to enhance the safety of the newly developed product.
TL;DR: In this article , prebiotic fructooligosaccharides (FOS) were extracted from garlic powder using ultrasound-assisted extraction (UAE), and the response surface methodology (RSM) was used to optimize the independent sonication variables, i.e., extraction temperature (ET, 80, 90, and 100 °C), amplitude level (AL, 70, 80 and 90%) and sonication time (ST, 10, 15 and 20 min).
Abstract: Dietary interventions have captured the attention of nutritionists due to their health-promoting aspects, in addition to medications. In this connection, supplementation of nutraceuticals is considered as a rational approach to alleviating various metabolic disorders. Among novel strategies, prebiotic-supplemented foods are an encouraging trend in addressing the issue. In the present investigation, prebiotic fructooligosaccharides (FOS) were extracted from garlic (Allium sativum L.) powder using ultrasound-assisted extraction (UAE). The response surface methodology (RSM) was used to optimize the independent sonication variables, i.e., extraction temperature (ET, 80, 90, and 100 °C), amplitude level (AL, 70, 80, and 90%) and sonication time (ST, 10, 15 and 20 min). The maximum FOS yield (6.23 ± 0.52%) was obtained at sonication conditions of ET (80 °C), AL (80%) and ST (10 min), while the minimum yield of FOS was obtained at high operating temperatures and time. The optimized FOS yield (7.19%) was obtained at ET (80 °C), AL (73%) and ST (15 min) after model validation. The influence of sonication parameters, i.e., ET, AL and ST, on FOS yield was evaluated by varying their coded levels from −1 to +1, respectively, for each independent variable. High-performance liquid chromatography with refractive index detector (HPLC-RID) detection and quantification indicated that sucrose was present in high amounts (2.06 ± 0.10 g/100 g) followed by fructose and glucose. Total FOS fractions which included nystose present in maximum concentration (526 ± 14.7 mg/100 g), followed by 1-kestose (428 ± 19.5 mg/100 g) and fructosylnystoses (195 ± 6.89 mg/100 g). Conclusively, garlic is a good source of potential prebiotics FOS and they can be extracted using optimized sonication parameters using ultrasound-assisted techniques with maximum yield percentage.