A novel peptide derived from Zingiber cassumunar rhizomes exhibits anticancer activity against the colon adenocarcinoma cells (Caco-2) via the induction of intrinsic apoptosis signaling

• Published in: PloS one 2024-06, Vol.19 (6), p.e0304701
• Main Authors: Promsut, Kitjasit, Sangtanoo, Papassara, Srimongkol, Piroonporn, Saisavoey, Tanatorn, Puthong, Songchan, Buakeaw, Anumart, Reamtong, Onrapak, Nutho, Bodee, Karnchanatat, Aphichart
• Format: Article
• Language: English
• Subjects: Adenocarcinoma; Adenocarcinoma - drug therapy; Adenocarcinoma - metabolism; Adenocarcinoma - pathology; Allergies; Annexin V; Antineoplastic Agents - chemistry; Antineoplastic Agents - pharmacology; Antioxidants; Antitumor activity; Apoptosis; Apoptosis - drug effects; Aquatic plants; Bcl-2 protein; Bcl-x protein; Biological activity; Biological properties; Biology and Life Sciences; Caco-2 Cells; Cancer; Care and treatment; Caspase-3; Colon; Colon cancer; Colonic Neoplasms - drug therapy; Colonic Neoplasms - metabolism; Colonic Neoplasms - pathology; Colorectal cancer; Digestive system; Electron spin; Enzymes; Exhibitions; Fractionation; Free Radical Scavengers - chemistry; Free Radical Scavengers - pharmacology; Free radicals; Functional foods; Functional foods & nutraceuticals; Gastrointestinal tract; Glycine; High performance liquid chromatography; Humans; Hydrolysates; Liquid chromatography; Mass spectrometry; Mass spectroscopy; Mcl-1 protein; Medicine and Health Sciences; Molecular docking; Molecular weight; mRNA; Oxidative stress; Pancreatin; Pepsin; Peptides; Peptides - chemistry; Peptides - pharmacology; Physical Sciences; Physiology; Polyimide resins; Polymerase chain reaction; Potassium; Protein hydrolysates; Proteins; Proteolysis; Quadrupoles; Real time; Recovery (Medical); Research and Analysis Methods; Rhizome - chemistry; Rhizomes; Scavenging; Scientific equipment and supplies industry; Signal Transduction - drug effects; Sodium; Solubility; Synthetic peptides; Toxicity; Zingiber cassumunar; Zingiberaceae - chemistry
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0304701

This paper presents the initial exploration of the free radical scavenging capabilities of peptides derived from protein hydrolysates (PPH) obtained from Zingiber cassumunar rhizomes (Phlai). To replicate the conditions of gastrointestinal digestion, a combination of pepsin and pancreatin proteolysis was employed to generate these hydrolysates. Subsequently, the hydrolysate underwent fractionation using molecular weight cut-off membranes at 10, 5, 3, and 0.65 kDa. The fraction with a molecular weight less than 0.65 kDa exhibited the highest levels ABTS, DPPH, FRAP, and NO radical scavenging activity. Following this, RP-HPLC was used to further separate the fraction with a molecular weight less than 0.65 kDa into three sub-fractions. Among these, the F5 sub-fraction displayed the most prominent radical-scavenging properties. De novo peptide sequencing via quadrupole-time-of-flight-electron spin induction-mass spectrometry identified a pair of novel peptides: Asp-Gly-Ile-Phe-Val-Leu-Asn-Tyr (DGIFVLNY or DY-8) and Ile-Pro-Thr-Asp-Glu-Lys (IPTDEK or IK-6). Database analysis confirmed various properties, including biological activity, toxicity, hydrophilicity, solubility, and potential allergy concerns. Furthermore, when tested on the human adenocarcinoma colon (Caco-2) cell line, two synthetic peptides demonstrated cellular antioxidant activity in a concentration-dependent manner. These peptides were also assessed using the FITC Annexin V apoptosis detection kit with PI, confirming the induction of apoptosis. Notably, the DY-8 peptide induced apoptosis, upregulated mRNA levels of caspase-3, -8, and -9, and downregulated Bcl-2, as confirmed by real-time quantitative polymerase chain reaction (RT-qPCR). Western blot analysis indicated increased pro-apoptotic Bax expression and decreased anti-apoptotic Bcl-2 expression in Caco-2 cells exposed to the DY-8 peptide. Molecular docking analysis revealed that the DY-8 peptide exhibited binding affinity with Bcl-2, Bcl-xL, and Mcl-1, suggesting potential utility in combating colon cancer as functional food ingredients.