Large numbers of bioactive peptides with potential applications in protecting against human diseases have been identified from plant sources. In this review, we summarized recent progress in the research of plant-derived bioactive peptides, encompassing their production, biological effects, and mechanisms. This review focuses on antioxidant, antimicrobial, antidiabetic, and anticancer peptides, giving special attention to evidence derived from cellular and animal models. Studies investigating peptides with known sequences and well-characterized peptidic fractions or protein hydrolysates will be discussed. The use of molecular docking tools to elucidate inter-molecular interactions between bioactive peptides and target proteins is highlighted. In conclusion, the accumulating evidence from in silico, in vitro and in vivo studies to date supports the envisioned applications of plant peptides as natural antioxidants as well as health-promoting agents. Notwithstanding, much work is still required before the envisioned applications of plant peptides can be realized. To this end, future researches for addressing current gaps were proposed.
The objective of this study is to establish conditions that allow optimal yield and antioxidant
activity for Golden Apple Snail (GAS) (Pomacea canaliculata) protein hydrolysate by employing
response surface methodology (RSM). A three level, face-centered, central composite design
(CCD) was adapted to assess the effects of temperature (45–65˚C); pH (8–10); the ratio of
enzyme to substrate (2–4%); and hydrolysis time (60–180 min). The antioxidative activity
of the hydrolysate obtained under optimized conditions was then evaluated via the following
metrics: hydroxyl radical scavenging, reducing power, and chelating effects on ferrous ion.
Established optimal conditions for the enzymatic protein hydrolysis of GAS were a temperature
of 45˚C, a pH of 10, an enzyme concentration of 2%, and hydrolysis time of 159 minutes. The
optimized GAS protein hydrolysate produced an experimental yield of 9.72% and antioxidant
activity of 73.54%—slightly less than the predicted yield of 11.36% and antioxidant activity of
78.88%. The optimized GAS protein hydrolysate formed demonstrated both higher chelating
effects and hydroxyl scavenging activity but had lower reducing power. These results suggest
that GAS protein hydrolysate holds potential as a natural antioxidant for use in food processing.
Utilization of palm kernel expeller (PKE), a palm oil milling by-product, may be diversified through the exploitation of its protein component. The PKE protein could be effectively extracted using an alkaline
solution and followed by enzymatic hydrolysis to produce PKE protein hydrolysates or crude PKE peptide. The extraction of PKE protein was successfully carried out using an alkaline solution at pH11, at ratio of 1:10 (g/ml), PKE powder to alkaline solution with continuous shaking, 150 rpm, in a water bath operating at 50°C for 30 min. The extracted protein powder (PKEP) had 68.50±3.08% crude protein, 0.54±0.03% fat and 0.73±0.02% ash. The freeze-dried PKEP was re-suspend in particular buffer and hydrolyzed with proteolytic enzymes (Alcalase® 2.4L, Flavourzyme® 500MG, pepsin or trypsin) to obtain PKEP hydrolysate (PKEPH). The effect of enzyme concentration (0, 2, 4, 6, 8 & 10%) and time of hydrolysis (0, 6, 12, 24, 48 h) was studied to determine the most efficient hydrolytic conditions. Results showed that all enzymes tested were capable of hydrolyzing the PKEP and producing hydrolysates with different degree of hydrolysis (DH%). At 8.0% concentration, Alcalase®2.4L hydrolyzed PKEP into the highest DH (75.96%) hydrolysate (PKEPH) after 1h hydrolysis. Although only with 2.0% Alcalase 2.4 L concentration, it was sufficient to produce PKEP hydrolysate of 81.35% DH %, but it required 12 h to hydrolyze the protein. Pepsin was relatively the least efficient protease to hydrolyze the PKEP.
Protein-rich by-products from the canning industry, especially dark flesh of skipjack, have limited uses due to several factors such as darken color, susceptibility to oxidation and off flavour. Protein hydrolysates from skipjack dark flesh was produced with different type of industrial proteases (Alcalase®2.4L FG, Protamex®, Neutrase®1.5MG and Flavourzyme®500MG) for 60, 120, 180 and 240 min with level of proteases used of 0.5, 1, 1.5 and 2% per weight of raw material. The degree of hydrolysis and free tryptophan content of hydrolysate were investigated. The results shows longer time with higher concentration of enzyme has increased the degree of hydrolysis. Alcalase®2.4L FG had the highest degree of hydrolysis among all proteases followed by Protamex®, Flavourzyme®500MG and Neutrase® 1.5MG. All enzymes increase free tryptophan content linearly with the increament of protease enzyme level. The longer the hydrolysis time, the higher the content of free tryptophan produced.
Cow (CwC) and camel casein (CaC) hydrolysates were generated using Alcalase™ (CwCA and CaCA) and Pronase-E (CwCP and CaCP) each for 3 and 6 h, and investigated for their potential to inhibit key lipid digesting enzymes i.e., pancreatic lipase (PL) and cholesteryl esterase (CE). Results revealed stronger PL and CE inhibition by CaC hydrolysates compared to CwC. Potent hydrolysates (CwCP-3 h and CaCA-6 h) upon simulated gastrointestinal digestion (SGID) showed significant improvement in inhibition of both PL and CE. However, both the SGID hydrolysates showed similar extent of PL and CE inhibition and were further sequenced for peptide identification. Peptides MMML, FDML, HLPGRG from CwC and AAGF, MSNYF, FLWPEYGAL from CaC hydrolysates were predicted to be most active PL inhibitory peptides. Peptide LP found in both CwC and CaC hydrolysates was predicted as active CE inhibitor. Thus, CwC and CaC could be potential source of peptides with promising CE and PL inhibitory properties.
Novel antihypercholesterolemic bioactive peptides (BAP) from peptic camel whey protein hydrolysates (CWPH) were generated at different time, temperature, and enzyme concentration (%). Hydrolysates showed higher pancreatic lipase- (PL; except 3 CWPH) and cholesterol esterase (CE)-inhibiting potential, as depicted by lower half-maximal inhibitory concentration values (IC50 values) compared with nonhydrolyzed camel whey proteins (CWP). Peptide sequencing and in silico data depicted that most BAP from CWPH could bind active site of PL, whereas as only 3 peptides could bind the active site of CE. Based on higher number of reactive residues in the BAP and greater number of substrate binding sites, FCCLGPVPP was identified as a potential CE-inhibitory peptide, and PAGNFLPPVAAAPVM, MLPLMLPFTMGY, and LRFPL were identified as PL inhibitors. Molecular docking of selected peptides showed hydrophilic and hydrophobic interactions between peptides and target enzymes. Thus, peptides derived from CWPH warrant further investigation as potential candidates for adjunct therapy for hypercholesterolemia.
Murrel namely Channa striatus or haruan contains all essential elements to maintain good health and to recover the lost energy after prolonged illness. The fatty acid composition (% of total fatty acid) indicated the abundant presence of C16:0 fatty acid as 30% and the other major fatty acids were C22:6 (15%), C20:4 (19%), C18:1 (12%) and C18:0 (15%). Haruan contains arachidonic acid (C20:4) as 19.0%, a precursor for prostaglandin and thromboxane biosyntheses. Both fatty and amino acids are important components for wound healing processes. Both the fillet and mucus extracts of haruan were found to exhibit a concentration dependent antinociceptive activity. In vitro antioxidant activity was higher in Channa roe protein hydrolysate than in Labeo roe protein hydrolysate in both DPPH radical scavenging and ferric reducing power. Protein content of roe concentrates (RPC) was found to be 90.2% (Channa) and 82.5% (Lates). Water absorption, oil absorption, foam capacity, stability and emulsifying capacity were found to be higher in Channa RPC than in Lates RPC. Characterization of protein hydrolysates from muscle and myofibrillar samples of haruan showed different kinetic and proteolytic activities. The skin extract of haruan influences the serotonergic receptor system thus they can function as an anti-depressant. Thus, haruan is the best example for food as medicine.
Modern agriculture prioritizes eco-friendly and sustainable strategies to enhance crop growth and productivity. The utilization of protein hydrolysate extracted from chicken feather waste as a plant biostimulant paves the path to waste recycling. A greenhouse experiment was performed to evaluate the implications of different doses (0, 1, 2, and 3 g L-1) of chicken feather protein hydrolysate (CFPH), application method (soil and foliar), and fertilizer rate (50% and 100%) on the growth performance of tea nursery plants. The highest dose of CFPH (3 g L-1) increased the shoot and root dry weights by 43% and 70%, respectively over control. However, no significant differences were observed between 2 and 3 g L-1 doses in plant dry weight, biometric, and root morphological parameters. Foliar application of CFPH significantly increased all the growth parameters compared to soil drenching except N, P, and K concentrations in leaves and roots. Plants grown under 100% fertilizer rate showed better growth performance than 50% fertilizer rate. Tea nursery plants treated with foliar 2 g L-1 dose and grown under full fertilizer rate recorded the highest plant dry weight, root length, and root surface area. However, tea plants under 50% fertilizer rate and treated with foliar 2 and 3 g L-1 doses sustained the growth similar to untreated plants under 100% fertilizer rate. The significantly higher N, P, and K concentrations in leaves were observed in plants treated with soil drenching of 2 and 3 g L-1 CFPH doses under 100% fertilizer rate. Our results indicate that the application of CFPH as a foliar spray is highly effective in producing vigorous tea nursery plants suitable for field planting, eventually capable of withstanding stress and higher yield.
Novel bioactive peptides from camel milk protein hydrolysates (CMPH) were identified and tested for inhibition of cholesterol esterase (CEase), and their possible binding mechanisms were elucidated by molecular docking. Papain-generated CMPH showed the highest degree of hydrolysis. All CMPH produced upon enzymatic degradation demonstrated a dramatic enhancement of CEase inhibition compared with intact camel milk proteins, with papain-generated hydrolysate P9 displaying the highest inhibition. Peptide identification and their modeling through PepSite 2 revealed that among 20 potential bioactive peptides in alcalase-generated hydrolysate A9, only 3 peptides, with sequences KFQWGY, SQDWSFY, and YWYPPQ, showed the highest binding toward CEase catalytic sites. Among 43 peptides in 9-h papain-generated hydrolysate P9, 4 peptides were found to be potent CEase inhibitors. Molecular docking revealed that WPMLQPKVM, CLSPLQMR, MYQQWKFL, and CLSPLQFR from P9 hydrolysates were able to bind to the active site of CEase with good docking scores and molecular mechanics-generalized born surface area binding energies. Overall, this is the first study reporting CEase inhibitory potential of peptides generated from milk proteins.
Matched MeSH terms: Protein Hydrolysates/chemistry
Actinopyga lecanora (A. lecanora) is classified among the edible species of sea cucumber, known to be rich in protein. Its hydrolysates were reported to contain relatively high antioxidant activity. Antioxidants are one of the essential properties in cosmeceutical products especially to alleviate skin aging. In the present study, pH, reaction temperature, reaction time and enzyme/substrate ratio (E/S) have been identified as the parameters in the papain enzymatic hydrolysis of A. lecanora. The degree of hydrolysis (DH) with antioxidant activities of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric-reducing antioxidant power (FRAP) assays were used as the responses in the optimization. Analysis of variance (ANOVA), normal plot of residuals and 3D contour plots were evaluated to study the effects and interactions between parameters. The best conditions selected from the optimization were at pH 5.00, 70 °C of reaction temperature, 9 h of hydrolysis time and 1.00% enzyme/substrate (E/S) ratio, with the hydrolysates having 51.90% of DH, 42.70% of DPPH activity and 109.90 Fe2+μg/mL of FRAP activity. The A. lecanora hydrolysates (ALH) showed a high amount of hydrophobic amino acids (286.40 mg/g sample) that might be responsible for antioxidant and antityrosinase activities. Scanning electron microscopy (SEM) image of ALH shows smooth structures with pores. Antityrosinase activity of ALH exhibited inhibition of 31.50% for L-tyrosine substrate and 25.40% for L-DOPA substrate. This condition suggests that the optimized ALH acquired has the potential to be used as a bioactive ingredient for cosmeceutical applications.
Matched MeSH terms: Protein Hydrolysates/pharmacology*
Jackfruit is a sweet tropical fruit with very pleasant aroma, and the ripe seeds are edible. In this study, jackfruit seed proteins were isolated and subjected to trypsin digestion. The resultant protein hydrolysate was then subjected to antioxidant assay-guided purification, using centrifugal filtration, C18 reverse-phase and strong cation exchange (SCX) fractionations. The purified SCX fraction was further analyzed by de novo peptide sequencing, and two peptide sequences were identified and synthesized. Peptide JFS-2 (VGPWQK) was detected with antioxidant potential, with EC50 value comparable to that of commercial GSH antioxidant peptide. Additionally, the identified peptides were tested with protein protection potential, in an albumin protein denaturation inhibitory assay. Concurrently, we also investigated the pH, temperature, and gastrointestinal-digestion stability profiles for the identified peptide. With further research efforts, the identified peptides could potentially be developed into preservative agent for protein-rich food systems or as health-promoting diet supplements.
Matched MeSH terms: Protein Hydrolysates/chemistry*
The stone fish is an under-utilized sea cucumber with many nutritional and ethno-medicinal values. This study aimed to establish the conditions for its optimum hydrolysis with bromelain to generate angiotensin I-converting enzyme (ACE)-inhibitory hydrolysates. Response surface methodology (RSM) based on a central composite design was used to model and optimize the degree of hydrolysis (DH) and ACE-inhibitory activity. Process conditions including pH (4-7), temperature (40-70 °C), enzyme/substrate (E/S) ratio (0.5%-2%) and time (30-360 min) were used. A pH of 7.0, temperature of 40 °C, E/S ratio of 2% and time of 240 min were determined using a response surface model as the optimum levels to obtain the maximum ACE-inhibitory activity of 84.26% at 44.59% degree of hydrolysis. Hence, RSM can serve as an effective approach in the design of experiments to improve the antihypertensive effect of stone fish hydrolysates, which can thus be used as a value-added ingredient for various applications in the functional foods industries.
Matched MeSH terms: Protein Hydrolysates/chemistry*
By-products from different animal sources are currently being utilised for beneficial purposes. Chicken processing plants all over the world generate large amount of solid by-products in form of heads, legs, bones, viscera and feather. These wastes are often processed into livestock feed, fertilizers and pet foods or totally discarded. Inappropriate disposal of these wastes causes environmental pollution, diseases and loss of useful biological resources like protein, enzymes and lipids. Utilisation methods that make use of these biological components for producing value added products rather than the direct use of the actual waste material might be another viable option for dealing with these wastes. This line of thought has consequently led to researches on these wastes as sources of protein hydrolysates, enzymes and polyunsaturated fatty acids. Due to the multi-applications of protein hydrolysates in various branches of science and industry, and the large body of literature reporting the conversion of animal wastes to hydrolysates, a large section of this review was devoted to this subject. Thus, this review reports the known functional and bioactive properties of hydrolysates derived from chicken by-products as well their utilisation as source of peptone in microbiological media. Methods of producing these hydrolysates including their microbiological safety are discussed. Based on the few references available in the literature, the potential of some chicken by-product as sources of proteases and polyunsaturated fatty acids are pointed out along with some other future applications.
As a protein-rich, underutilized crop, green soybean could be exploited to produce hydrolysates containing angiotensin-I converting enzyme (ACE) inhibitory peptides. Defatted green soybean was hydrolyzed using four different food-grade proteases (Alcalase, Papain, Flavourzyme and Bromelain) and their ACE inhibitory activities were evaluated. The Alcalase-generated green soybean hydrolysate showed the highest ACE inhibitory activity (IC50: 0.14 mg/mL at 6 h hydrolysis time) followed by Papain (IC50: 0.20 mg/mL at 5 h hydrolysis time), Bromelain (IC50: 0.36 mg/mL at 6 h hydrolysis time) and Flavourzyme (IC50: 1.14 mg/mL at 6 h hydrolysis time) hydrolysates. The Alcalase-generated hydrolysate was profiled based on its hydrophobicity and isoelectric point using reversed phase high performance liquid chromatography (RP-HPLC) and isoelectric point focusing (IEF) fractionators. The Alcalase-generated green soybean hydrolysate comprising of peptides EAQRLLF, PSLRSYLAE, PDRSIHGRQLAE, FITAFR and RGQVLS, revealed the highest ACE inhibitory activity of 94.19%, 99.31%, 92.92%, 101.51% and 90.40%, respectively, while their IC50 values were 878 μM, 532 μM, 1552 μM, 1342 μM and 993 μM, respectively. It can be concluded that Alcalase-digested green soybean hydrolysates could be exploited as a source of peptides to be incorporated into functional foods with antihypertensive activity.
Matched MeSH terms: Protein Hydrolysates/isolation & purification; Protein Hydrolysates/pharmacology*
Palm kernel cake protein was hydrolyzed with different proteases namely papain, bromelain, subtilisin, flavourzyme, trypsin, chymotrypsin, and pepsin to generate different protein hydrolysates. Peptide content and iron-chelating activity of each hydrolysate were evaluated using O-phthaldialdehyde-based spectrophotometric method and ferrozine-based colorimetric assay, respectively. The results revealed a positive correlation between peptide contents and iron-chelating activities of the protein hydrolysates. Protein hydrolysate generated by papain exhibited the highest peptide content of 10.5 mM and highest iron-chelating activity of 64.8% compared with the other hydrolysates. Profiling of the papain-generated hydrolysate by reverse phase high performance liquid chromatography fractionation indicated a direct association between peptide content and iron-chelating activity in most of the fractions. Further fractionation using isoelectric focusing also revealed that protein hydrolysate with basic and neutral isoelectric point (pI) had the highest iron-chelating activity, although a few fractions in the acidic range also exhibited good metal chelating potential. After identification and synthesis of papain-generated peptides, GGIF and YLLLK showed among the highest iron-chelating activities of 56% and 53%, whereas their IC50 were 1.4 and 0.2 μM, respectively.
Matched MeSH terms: Protein Hydrolysates/pharmacology*; Protein Hydrolysates/chemistry
Protein hydrolysates produced from different food sources exhibit therapeutic potential and can be used in the management of chronic diseases. This study was targeted to optimise the conditions for the hydrolysis of stone fish protein to produce antioxidant hydrolysates using central composite design (CCD) by response surface methodology (RSM). The stone fish protein was hydrolysed under the optimum predicted conditions defined by pH (6.5), temperature (54°C), E/S ratio (1.5%), and hydrolysis time (360 min). The hydrolysates were then evaluated for 2,2-diphenyl-1-picrylhydrazyl radical (DPPH•) scavenging activity and ferrous ion- (Fe2+-) chelating activity. Results validation showed no significant difference between the experimental values of DPPH• scavenging activity (48.94%) and Fe2+-chelating activity (25.12%) obtained at 54.62% degree of hydrolysis (DH) compared to their corresponding predicted values of 49.79% and 24.08% at 53.08% DH, respectively. The hydrolysates demonstrated non-Newtonian behavior (n < 1) with stronger shear-thinning effect and higher viscosities at increasing concentration. Thus, RSM can be considered as a promising strategy to optimise the production of stone fish protein hydrolysates containing antioxidant peptides. It is hoped that this finding will enhance the potential of stone fish protein hydrolysates (SHs) as therapeutic bioactive ingredient in functional foods development.
The emergence of bacterial resistance against conventional antibiotics and the growing interest in developing alternative, natural antibacterial agents have prompted the search for plant-derived antibacterial peptides in recent decades. Different classes of endogenous antibacterial peptides have been identified from various plant species. Moreover, protein hydrolysates and hydrolysate-derived peptides with potent antibacterial effects have also been identified from numerous plant sources. Antibacterial peptides are often cationic and amphipathic, consisting of fewer than 100 amino acids. They are able to disrupt bacterial membrane integrity via pore formation and/or compromise bacterial metabolic processes. In this review, we summarize current knowledge on the characteristics and modes of action of antibacterial peptides, as well as salient points concerning the production of antibacterial protein hydrolysates from plant proteins. Examples of plant-derived antibacterial hydrolysates and peptides will be highlighted, with particular attention to less explored seeds, fermented plant foods and agricultural by-products. Promising future research directions with regards to the application of plant-derived antibacterial hydrolysates and peptides in food preservation, farm animal disease management, and nutraceutical/functional food development will be proposed.
This study was conducted to evaluate umami taste in protein hydrolysate produced from green mussel (Perna viridis) by hydrolysing with flavorzyme at pH 8, enzyme substrate ratio (E/S) 3% with or without the presence of 0.4% sodium tripolyphosphate (STPP) and 1.5% NaCI. Degree of hydrolysis (DH), yield, amino acid compositions, molecular weight distribution and sensory evaluation were determined. The highest DH (23.18%), darkest color and highest yield (8.34%) were recorded for hydrolysate produced in the presence of both STPP and NaCI. Electrophoresis analysis showed the presence of protein bands between 10 to 70 kDa where hydrolysate with addition of STPP and NaCI had bands with lower intensities. Amino acids which contribute to the umami taste such as glutamic acid, glycine and aspartic acid were higher in hydrolysate produced with STPP and NaCI addition. The hydrolysate has lesser fishy odor and flavor than those produced with only in the presence of flavorzyme and was also rated with highest score for all the five basic tastes including bitterness. However, the score for bitterness was still lower than the reference solutions. Therefore, green mussel hydrolysate produced in this study has a good potential as a food flavorant.
Tilapia is a popular freshwater fish and among the important cultured fish grown worldwide. In this study, fish protein
hydrolysate was produced from tilapia (Oreochromis niloticus) by-product (TB) and tilapia muscle (TM) through enzymatic
hydrolysis using alcalase. The TB and TM protein hydrolysates were evaluated for its characteristics in terms of angiotensin
I-converting enzyme (ACE) inhibition activity, peptide size distribution, and functional properties. Hydrolysis for 1 h for
TB and TM successfully produced low molecular weight peptides (<14.2kDa) with the highest ACE inhibitory activities.
The findings also demonstrated that both samples have high nitrogen solubility (>80% at pH2-9) and good emulsifying,
water and oil holding capacities. The study indicated that tilapia protein hydrolysates have the potential to be used as
functional food products.
Threadfin bream (Nemipterus japonicas) muscle was hydrolysed using protease extracted from
bilimbi (Averrhoa bilimbi L.) fruit. This study was performed in order to compare the efficiency of bilimbi protease in producing threadfin bream protein hydrolysate with the commercial protease; alcalase 2.4 L. Initially, protease was extracted and then purified using 40% ammonium sulfate precipitation method. The proteolytic activity of the crude extract and purified protease was determined. Precipitation using 40% ammonium sulfate resulted in bilimbi protease specific activity of 2.36 U/mg and 23.13% recovery. Threadfin bream hydrolysate was prepared based on the pH-stat method by hydrolysis for 2 hrs. Hydrolysis using bilimbi protease produced 34.76% degree of hydrolysis (DH) and 3.75% yield while hydrolysis using alcalase resulted in 86.6% DH with 22.78% yield. Alcalase hydrolysate showed higher solubility than bilimbi protease hydrolysate at pH 7 with 70.87 and 32.16% solubility, respectively. Results also showed that protein content of threadfin bream hydrolysate produced using alcalase was higher (86.86%) than those produced using bilimbi protease (22.12%). However, both hydrolysates showed low moisture content between 3.93 to 7.00%. The molecular weight distribution analysis using SDS–PAGE indicated the distribution of smaller peptides especially in alcalase hydrolysate. Overall, the results showed that alcalase is more efficient enzyme choice than bilimbi protease for preparing threadfin bream hydrolysates. However, both hydrolysates could play an important role thus contribute to the food industry.