Anti-Inflammatory and Anti-Nociceptive Activities of Native and Modified Hen Egg White Lysozyme

Wilman Carrillo,Humberto Spindola,Mercedes Ramos,Isidra Recio,Joao Ernesto Carvalho 한국식품영양과학회 2016 Journal of medicinal food Vol.19 No.10

Persistent inflammatory conditions can have severe pathological consequences. Although the use of nonsteroidal anti-inflammatory drugs (NSAIDs) is effective, it has side effects, particularly at the gastrointestinal level. There is then a high interest to identify natural anti-inflammatory compounds with no side effects. The anti-inflammatory and anti-nociceptive activities of hen egg lysozyme (LZ), both in its native form and modified by heat treatment, chemically or by enzymatic digestion have been tested in this study. The carrageenan-induced model in mice using native LZ or modified LZ has been applied. It was observed that LZ denatured by heat treatment at pH 6.0 presented 39.47% of inhibition of paw edema when administered at 30 mg/kg. LZ denatured with DL-dithiothreitol (DTT) presented a significant result of 42.10% inhibition of paw edema when administered at 30 mg/kg of animal weight. Modified LZ showed anti-inflammatory capacity comparable with the activity of the positive control dexamethasone. A classical model of acetic acid-induced abdominal writhing tests in mice was used to assess anti-nociceptive activity of native LZ and denatured heat treatment LZ and denatured chemical agent LZ. Finally, hydrolyzed native LZ presented 48% of inhibition of abdominal writhing in mice. Modified LZ with heat, chemical, and hydrolysis presented anti-inflammatory and anti-nociceptive activities independently of their natural enzymatic activity. These novel data point out the potential use of denatured and digested LZ as therapeutic agents and offer alternatives to the use of NSAIDs. LZ can be a natural source of anti-inflammatory and anti-nociceptive agents.

Antiproliferative Activity of Walnut (Juglans regia L.) Proteins and Walnut Protein Hydrolysates

Wilman Carrillo,Jose Angel Gomez-Ruiz,Ana Lucia Ruiz,Joao Ernesto Carvalho 한국식품영양과학회 2017 Journal of medicinal food Vol.20 No.11

Proteins from Juglans regia L. were isolated. Then, proteins were hydrolyzed with different enzymes. Antiproliferative activity of proteins and of the protein hydrolysates of J. regia L. were evaluated using the sulforhodamine B method. Glutelin and prolamin proteins presented a high antiproliferative activity against cancer cells PC-3 (prostate) and K-562 (leukemia) with values of 43.9 and 84.4 μg/mL, respectively. The highest inhibitory effect observed was 50% at 0.25 μg/mL concentration in gastrointestinal digestion with pepsin and corolase pp in a dose-dependent manner against cancer cell UACC62 (melanoma). Pepsin hydrolysate showed inhibitory effects against cancer cell UACC62 (melanoma) with a concentration of 71.0 μg/mL. The effects were studied in a dose-dependent manner. The hydrolysate obtained with neutrase enzyme presented inhibitory effects against cancer cell UACC62 (melanoma) at a concentration of 25 μg/mL. Neither proteins nor protein hydrolysates presented cytotoxicity against normal cell assay VERO (epithelial).

Identification of Antimicrobial Peptides of Native and Heated Hydrolysates from Hen Egg White Lysozyme

Wilman Carrillo,Mercedes Ramos 한국식품영양과학회 2018 Journal of medicinal food Vol.21 No.9

Hen eggs are a source of bioactive compounds, of which the hen egg white lysozyme (HEWL) protein. HEWL has a demonstrated antibacterial activity. The aim of this study was to evaluate the antimicrobial activity of native and heated HEWL hydrolysates obtained through hydrolysis with pepsin and to identify their peptides using the reversed phase high performance liquid chromatography–electrospray ionization–tandem mass spectrometry (RP-HPLC-ESI-MS-MS) analysis. Native and heat-treated HEWL was hydrolyzed with pepsin at pH 1.2, and their antibacterial activity was tested against Escherichia coli and Staphylococcus carnosus. Two of the hydrolysates obtained presented high antibacterial activity against Gram-positive and Gram-negative bacteria. Native HEWL hydrolysate was a bactericide at 2.0 mg/mL against E. coli. Fifty-one peptide sequences were identified on the two hydrolysates. Peptides identified are cationic peptides. These peptides are rich in Lys and Arg cationic amino acids and have Trp in their sequences.

Antiulcerative and Antinociceptive Activities of Casein and Whey Proteins

Wilman Carrillo,Karin Maia Monteiro,Humberto Spindola,Mercedes Ramos,Joao Ernesto de Carvalho 한국식품영양과학회 2018 Journal of medicinal food Vol.21 No.6

The aim of this study was to evaluate the antiulcerative and antinociceptive activities of milk proteins using the induced gastric ulcer with ethanol rat model and the acetic acid-induced writhing mouse model. Casein (CN), (100, 300, and 1000 mg kg−1) doses presented antiulcerative activity on a dose-dependent manner with values of 30.8%, 41.4%, and 57.0% of inhibition measured using the ulcerative lesions index (ULI), respectively. Whey protein concentrate (WPC), (100, 300, and 1000 mg kg−1) doses presented antiulcerative activity on a dose-dependent manner with values of 48.9%, 65.5%, and 68.22% of ULI inhibition, respectively. CN, casein hydrolysates (CNH), WPC, and whey protein hydrolysates (WPH), (3, 10, and 30 mg kg−1) doses presented antinociceptive activity using the acetic acid-induced writhing in the mouse model. CN (30 mg kg−1) presented a value of 40% of inhibition writhing, and CNH (30 mg kg−1) presented antinociceptive activity with a value up to 46% of writhing inhibition. WPC (30 mg kg−1) presented a value of 52.50%, and WPH (30 mg kg−1) presented antinociceptive activity with a value up to 88.00% of writhing inhibition. In conclusion, CN and WPC demonstrated in vivo antiulcerative properties and represent a promising alternative to be used as protectors of the gastric mucosa. CNH and WPH demonstrated in vivo antiulcerative properties and represent a promising alternative to be used as natural analgesic.

Antiulcerative Activity of Milk Proteins Hydrolysates

Wilman Carrillo,Karin Maia Monteiro,Daniel Martı´nez-Maqueda,Mercedes Ramos,Isidra Recio,Joa˜o Ernesto de Carvalho 한국식품영양과학회 2018 Journal of medicinal food Vol.21 No.4

Several studies have shown the protective effect of dairy products, especially α-lactalbumin and derived hydrolysates, against induced gastric ulcerative lesions. The mucus strengthening represents an important mechanism in the defense of gastrointestinal mucosa. Previously, a hydrolysate from casein (CNH) and a hydrolysate from whey protein concentrate rich in β-lactoglobulin (WPH) demonstrated a stimulatory activity on mucus production in intestinal goblet cells. The aim of this work was to evaluate the possible antiulcerative activity of these two hydrolysates in an ethanol-induced ulcer model in rats. All tested samples significantly reduced the ulcerative lesions index (ULI), compared with the saline solution, using doses of 300 and 1000 mg kg−1 body weight with decreases up to 66.3% ULI. A dose–response relationship was found for both hydrolysates. The involvement of endogenous sulfhydryl (SH) groups and prostaglandins (PGs) in the antiulcerative activity was evaluated using their blockage. The antiulcerative activity of WPH showed a drastic decrease in presence of N-ethylmaleimide (from 41.4% to 9.2% ULI). However, the CNH antiulcerative properties were not significantly affected. The cytoprotective effect of WPH appears to depend on a PG-mediated mechanism. In conclusion, CNH and WPH demonstrated in vivo antiulcerative properties and represent a promising alternative as protectors of the gastric mucosa.

Isolation of Antibacterial Hydrolysates from Hen Egg White Lysozyme and Identification of Antibacterial Peptides

Wilman Carrillo,Araceli Lucio,Juan Gaibor,Dayana Morales,Grace Vasquez 한국식품영양과학회 2018 Journal of medicinal food Vol.21 No.8

Native and heated hen egg white lysozyme (HEWL) hydrolysates were isolated by hydrolysis with pepsin at pH 2.0 in situ in a cation exchange membrane to isolate and identify antibacterial peptides of the HEWL hydrolysates. Native and heated HEWL was partially hydrolyzed with pepsin at pH 2.0. The fractions were eluted with 5 M ammonia to identify 23 antibacterial peptides using a tandem mass spectrometry. Then, these fractions were eluted with a solution of NaCl 1 M, and seven positively charged peptides f(23–28) YSLGNW, f(122–129) AWIRGCRL, f(123–129) WIRGCRL, f(124–129) IRGCRL, f(82–96) ALLSSDITASVNCAK, f(103–129) VAWRNRCKGTDVQAWIRGCRL, and f(97–123) KIVSDGNGMNAWVAWRNRCKGT were identified using tandem mass spectrometry. Native HEWL hydrolysate presented an enzymatic activity of 23.0%, heated HEWL hydrolysate at pH 6.0 presented a residual enzymatic activity of 22.0%, and heated HEWL hydrolysate at pH 7.0 presented an enzymatic activity of 21.33%. Native and heated HEWL hydrolysate presented antibacterial activity against Escherichia coli and Staphylococcus carnosus. Native HEWL hydrolysate presented a higher enzymatic activity than heated HEWL hydrolysates.