Purification, Identification and Characterization of Antioxidant Peptides from Corn Silk Tryptic Hydrolysate: An Integrated In Vitro-In Silico Approach

Corn silk (CS) is an agro-by-product from corn cultivation. It is used in folk medicines in some countries, besides being commercialized as health-promoting supplements and beverages. Unlike CS-derived natural products, their bioactive peptides, particularly antioxidant peptides, are understudied. This study aimed to purify, identify and characterize antioxidant peptides from trypsin-hydrolyzed CS proteins. Purification was accomplished by membrane ultrafiltration, gel filtration chromatography, and strong-cation-exchange solid-phase extraction, guided by 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt radical cation (ABTS•+) scavenging, hydrogen peroxide scavenging, and lipid peroxidation inhibition assays. De novo sequencing identified 29 peptides (6–14 residues; 633–1518 Da). The peptides consisted of 33–86% hydrophobic and 10–67% basic residues. Molecular docking found MCFHHHFHK, VHFNKGKKR, and PVVWAAKR having the strongest affinity (−4.7 to −4.8 kcal/mol) to ABTS•+, via hydrogen bonds and hydrophobic interactions. Potential cellular mechanisms of the peptides were supported by their interactions with modulators of intracellular oxidant status: Kelch-like ECH-associated protein 1, myeloperoxidase, and xanthine oxidase. NDGPSR (Asn-Asp-Gly-Pro-Ser-Arg), the most promising peptide, showed stable binding to all three cellular targets, besides exhibiting low toxicity, low allergenicity, and cell-penetrating potential. Overall, CS peptides have potential application as natural antioxidant additives and functional food ingredients.

Comparative Assay of the Nutritional Composition and Antioxidant Effect of the Cotyledon and Pulp of Chrysophyllum albidum fruit

Background: Chrysophyllum albidum has been widely consumed for its flesh part as a fruit snack and source of vitamins but also grossly under-utilized because of dearth of knowledge on nutritional and therapeutic potencies of other fruit parts. This study thus aimed to comparatively determine the nutritional, phytochemical and in vitro antioxidant properties of the flesh and cotyledon of C. albidum. Methods: Proximate and phytochemical contents were determined according to the methods of Association of Official Analytical Chemists (2000). Mineral concentrations were evaluated using Atom Analyzer according to the protocols of AOAC (2000). Antioxidant properties were assayed through the 2,2-diphenyl-1-picryl hydrazyl radical scavenging, reducing power and lipid peroxidation abilities according to the methods of Barros et al (2007). Results: Findings indicated a higher percentage contents of ash (3.83 ± 0.38), moisture (13.86 ± 0.84), crude fiber (11.07 ± 2.72) and crude protein (7.44 ± 0.44) in the flesh than the cotyledon of C. albidum, which were insignificantly different (p > 0.05). On the other hand, crude fat (13.80 ± 2.60) and total carbohydrate (64.96 ± 2.77) were found to be more in the cotyledon than the flesh but also not significantly different (p > 0.05). The mineral analysis revealed a higher but insignificantly different (p > 0.05) concentrations of iron (2.31 ± 0.22), copper (1.23 ± 0.09), zinc (2.94 ± 0.12) and potassium (1.48 ± 0.09) in the flesh than the cotyledon whereas cobalt (3.09 ± 0.92), magnesium (21.13 ± 0.58), sodium (16.27 ± 0.62) and selenium (4.24 ± 0.28) were more in the cotyledon than the flesh although insignificantly different (p > 0.05). While the flesh was observed to significantly contain high total phenol values, the cotyledon showed more significant mean values for tannin, oxalate, saponin and β-carotene than the flesh. The in vitro DPPH free radical scavenging and lipid peroxidation inhibition findings indicated higher antioxidant activities in the flesh than in the seed. Conclusion: Conclusively, the flesh and seed fruit parts of C. albidum showed considerable and significant amounts of the parameters under study, which can be further exploited for their nutritional and pharmacological essence.

The antioxidative-activity stability of aloe vera (Aloe vera var. chinensis) instant during storage

Aloe vera contains a phenolic compound that has bioactive activity. Previous research showed that microencapsulation of aloe vera powder with maltodextrin as an encapsulation agent produced instant aloe vera with high antioxidative activity. The problem was the hygroscopic instant caused rapid moisture and oxygen absorption during storage, therefore decreasing the instant aloe vera antioxidative activity periodically. The aim of this research was to evaluate the antioxidative activity stability of instant aloe vera during storage. The processing of instant aloe vera through a reconstituted aloe vera powder with water with a ratio of 1:120 and then added with 2.5% maltodextrin as the encapsulating agent. The solution was then inserted into a spray dryer with an inlet temperature of 130oC, an outlet temperature of 103oC, and the flow rate of the solution is 350.0 mL/h. The resulted instant aloe vera was divided into 15 packs with a weight of 25 g, and each sample was wrapped with polyethylene plastic film with 0.80 mm thickness and then was stored at 25oC with a relative humidity of 75%. The sample was conducted in triplicate. The moisture content, and antioxidative activity that was based on the ability to capture 1,1-diphenyl-2- picrylhydrazyl (DPPH) radical (RSA) and lipid peroxidation inhibition were analyzed every week until the critical condition was achieved at a moisture level of 12%. The research showed that the radical scavenging activity (RSA) and lipid peroxidation inhibition of instant aloe vera before storage were 16.34±1.22% and 39.33±1.68%, respectively, whereas in the critical condition the RSA was 3.63±0.04% and the lipid peroxidation inhibition was 22.31±0.02%. Based on their antioxidative activity, the appropriate storage time of instant aloe vera was about 12 weeks in polyethylene plastic film of 0.08 mm thickness

Effect of ethanolic extract of Silybum marianum L. Gaertn. on lipid peroxidation inhibition and microbial count in minced beef

The present research was carried out for two main targets, the first one is phytochemical and biological screening of the aerial parts of Silybum marianum. The second one is the possibility of using Silybum marianum ethanolic extract as natural preservative for extending minced beef shelf-life during refrigeration and frozen storage. The chemical analysis of dry matter revealed that total ash content was 13.04% of D.M, crude fibers, total phenolic compounds, total flavonoids and total tannins were (24.59%, 9.16%, 0.84% and (0.08%), respectively. The highest values of antioxidant activity were recorded in ethyl acetate and ethyl alcohol 70%extacts, respectively for successive and selective solvents with values of 82.45% and 58.85% for successive solvents, 86.84% and 70.50% for selective solvents, respectively. Antimicrobial activity showed that, S. marianum L. Gaertn. had effect on gram positive, gram negative bacteria and yeast but had no effect on different strains of studied fungi. S. marianum had an influence on microorganisms growth in minced beef, the highest effect was 1.0% for 60 days (56 CFU). The most effective concentrations on lipid peroxidation prevention in minced beef were 1.5% for 3 days and 1.5 for 60 days.

Purification, Identification, Activity Evaluation, and Stability of Antioxidant Peptides from Alcalase Hydrolysate of Antarctic Krill (Euphausia superba) Proteins

For utilizing the largest source of marine proteins, Antarctic krill (Euphausia superba) proteins were defatted and hydrolyzed separately using pepsin, alcalase, papain, trypsin, and netrase, and alcalase hydrolysate (EPAH) showed the highest DPPH radical (DPPH·) and hydroxyl radical (HO·) scavenging activity among five hydrolysates. Using ultrafiltration and chromatography methods, fifteen antioxidant peptides were purified from EPAH and identified as Asn-Gln-Met (NQM), Trp-Phe-Pro-Met (WFPM), Gln-Asn-Pro-Thr (QNPT), Tyr-Met-Asn-Phe (YMNF), Ser-Gly-Pro-Ala (SGPA), Ser-Leu-Pro-Tyr (SLPY), Gln-Tyr-Pro-Pro-Met-Gln-Tyr (QYPPMQY), Glu-Tyr-Glu-Ala (EYEA), Asn-Trp-Asp-Asp-Met-Arg-Ile-Val-Ala-Val (NWDDMRIVAV), Trp-Asp-Asp-Met-Glu-Arg-Leu-Val-Met-Ile (WDDMERLVMI), Asn-Trp-Asp-Asp-Met-Glu-Pro-Ser-Phe (NWD-DMEPSF), Asn-Gly-Pro-Asp-Pro-Arg-Pro-Ser-Gln-Gln (NGPDPRPSQQ), Ala-Phe-Leu-Trp-Asn (AFLWA), Asn-Val-Pro-Asp-Met (NVPDM), and Thr-Phe-Pro-Ile-Tyr-Asp-Tyr-Pro-Gln (TFPIYDPQ), respectively, using a protein sequencer and ESI/MS. Among fifteen antioxidant peptides, SLPY, QYPPMQY and EYEA showed the highest scavenging activities on DPPH· (EC50 values of 1.18 ± 0.036, 1.547 ± 0.150, and 1.372 ± 0.274 mg/mL, respectively), HO· (EC50 values of 0.826 ± 0.027, 1.022 ± 0.058, and 0.946 ± 0.011 mg/mL, respectively), and superoxide anion radical (EC50 values of 0.789 ± 0.079, 0.913 ± 0.007, and 0.793 ± 0.056 mg/mL, respectively). Moreover, SLPY, QYPPMQY and EYEA showed strong reducing power, protective capability against H2O2-damaged plasmid DNA, and lipid peroxidation inhibition ability. Furthermore, SLPY, QYPPMQY, and EYEA had high stability under temperatures lower than 80 °C, pH values ranged from 6–8, and simulated GI digestion for 180 min. The results showed that fifteen antioxidant peptides from alcalase hydrolysate of Antarctic krill proteins, especially SLPY, QYPPMQY and EYEA, might serve as effective antioxidant agents applied in food and health products.

Survey of Phenolic Acids, Flavonoids and In Vitro Antioxidant Potency Between Fig Peels and Pulps: Chemical and Chemometric Approach

In the present study, chromatic coordinates, phenolic acids, flavonoids and antioxidant capacity assessed by 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonate (ABTS) and lipid peroxidation inhibition capacity (LPIC) essays and their relative IC50 were investigated in 25 fig cultivars growing in Morocco. The aims of this study were to determine (i) the variation in these compounds among light and dark-colored cultivars, (ii) their partitioning between fruit peel and pulp and (iii) to display network connections among these variables. Twelve phenolic compounds (PCs) were isolated in peel extract versus eight in pulp samples. Anthocyanins, mainly cyanidin-3,5-diglucoside and cyanidin-3-O-rutinoside, were the predominant compounds in peels, where the mean concentrations were 75.90 ± 18.76 and 77.97 ± 18.95 µg/g dw, respectively. On the other hand, (−)-epicatechin and cyanidin-3-O-rutinoside were the major compounds in the pulp extracts, where the mean values were 5.23 ± 4.03 and 9.01 ± 5.67 µg/g dw, respectively. A two-dimensional hierarchically clustered heatmap was applied to the dataset to explore correlations in the dataset and similarities between cultivars, without dimensionality reduction. Results showed that anthocyanins, particularly pelargonidin-3-O-rutinoside, cyanidin-3,5-diglucoside and cyanidin-3-O-rutinoside, were the main contributors to the peels’ free radical scavenging capacity. This capacity was particularly higher in the peel of dark-colored figs compared to the fruit pulp. The local cultivar “INRA 1301” showed the most promising phenolic profile due to its very high levels of almost all detected PCs, especially (−)-epicatechin, quercetin-3-O-rutinoside, quercetin-3-O-glucoside, cyanidine-3,5-diglucoside, cyanidine-3-O-rutinoside and pelargonidin-3-O-rutinoside (54.66, 141.08, 35.48, 494.08, 478.66, 12.56 µg/g dw, respectively). Having the darkest figs in the collection (L* = 25.72, c* = 22.09 and h° = 20.99), this cultivar has also combined promising IC50 values, which were of 19.85, 40.58 and 124.78 µg/mL for DPPH, ABTS and LPIC essays, respectively.

Intracellular reactive oxygen species scavenging activity and lipid peroxidation inhibition by secondary metabolites isolated from the endophytic fungus, Daldinia eschscholtzii

Fungal endophytes associated with medicinal plants are of research interests, since they provide a viable alternative source for molecules with medicinal value. In this study, we report for the first time two fungal endophytic isolates, AI.EF 001 and AI.EF 002 belonging to the genus Daldinia from the leaves of medicinal plant, Abutilon indicum linn (AI). Both AI.EF were identified as Daldinia eschscholtzii (DE) species by ITS1-5.8-ITS2 sequence analysis and phylogenetic tree reconstruction by Neighbor-joining method. Crude extracts of DE (EFEA), generated by ethylacetate/water fractionation of the fungal methanol extracts when subjected to column chromatography separation yielded 5 compounds. NMR and other spectral data revealed the compounds to be ɑ-napthoflavone, Syringaldehyde, 3,4,5-trimethoxy benzoic acid, 2-Furoic acid and Gossypetin 3′ O glycoside. All of these compounds are being reported for the first time from DE. The isolated compounds showed promising free radical scavenging activities. The compounds also exhibited anti inflammatory property by down regulating intracellular ROS as well as inhibiting LPS induced lipid peroxidation in AtT 20 mouse pituitary cells. Current finding demonstrates endophyte DE as a new source for the flavanoid, Gossypetin-3′-O-glycoside along with other phytoconstituents with strong antioxidant and anti-inflammatory activity.

Effects of Cooking Method on the Antioxidant Activity and Inhibition of Lipid Peroxidation of the Javanese Salad “Pecel” Vegetables and Its Peanut Sauce Dressing

Vegetables are essential in our diet to maintain health, partly due to their antioxidant properties. A well-known Javanese salad called “Pecel” is prepared by boiling the vegetables and dressed with seasoned peanut sauce. Cooking can reduce or improve the antioxidant properties of foods; therefore, the purpose of this study was to evaluate the effects of brief water boiling (1 min), steaming (1 min), and water blanching (20 s) of the Javanese Pecel vegetables, with or without the peanut sauce. We assessed the in vitro antioxidant capacity and lipid peroxidation inhibition of the salad samples prepared using each cooking method. Six vegetables, i.e., Sesbania grandiflora (turi) flower, Amaranthus hybridus L. (spinach), Carica papaya (papaya) leaves, Cosmos caudatus L. (kenikir) leaves, Vigna unguiculata ssp. Sesquipedalis (yard-long beans), and Vigna radiata (mung-bean) sprouts were cooked by boiling or steaming for 1 min or blanching for 20 s. Peanut (Arachis hypogaea), the raw material for peanut sauce, was fried in either fresh palm oil or repeatedly used palm oil. Our results revealed that the highest antioxidant capacity (percent inhibition of DPPH radicals) was observed following boiling for 1 min in case of spinach ( 41.94 ± 9.8 %), papaya ( 59.04 ± 5.35 %), kenikir ( 54.93 % ± 6.32 % ), and yard-long beans ( 70.21 ± 8.91 %); steaming for 1 min in case of turi flower ( 60.25 ± 3.63 %); and blanching for 20 s in case of mung-bean sprouts ( 49.27 ± 3.69 %). Peanut sauce prepared by frying peanuts in fresh or repeatedly used palm oil reduces the natural antioxidant and lipid peroxidation inhibition properties. However, seasoning the peanut sauce with fresh garlic and lime leaves can restore the lost antioxidant properties. Our study provides the first and clear evidence of the optimal cooking method for Pecel vegetables and sheds light on the wisdom behind the existing traditional cooking method.