Semisynthesis and biological evaluation of novel honokiol thioethers against colon cancer cells HCT116 via inhibiting the transcription and expression of YAP protein.

Honokiol, derived from Magnolia officinalis (a traditional Chinese medicine), has been reported to have anticancer activity. Here, a series of novel honokiol thioethers bearing a 1,3,4-oxadiazole moiety were prepared and evaluated for their anticancer activities against three types of digestive system tumor cells. Biological evaluation showed that honokiol derivative 3k exhibited the best antiproliferative activity against HCT116 cells with an IC50 value of 6.1 µmol/L, superior to the reference drug 5-fluorouracil (IC50: 9.63 ± 0.27 µmol/L). The structure-activity relationships (SARs) indicated that the introduction of -(4-NO2)Ph, 3-pyridyl, -(2-F)Ph, -(4-F)Ph, -(3-F)Ph, -(4-Cl)Ph, and -(3-Cl)Ph groups was favorable for enhancing the anticancer activity of the title honokiol thioethers. Further study revealed that honokiol thioether 3k can well inhibit the proliferation of colon cancer cells HCT116, arresting the cells in G1 phase and inducing cell death. Moreover, a preliminary mechanism study indicated that 3k directly inhibits the transcription and expression of YAP protein without activating the Hippo signaling pathway. Thus, honokiol thioether 3k could be deeply developed for the development of honokiol-based anticancer candidates.

Identification and Characterization of Novel Umami Peptides from Protein Hydrolysates of Morchella esculenta and Their Interaction with T1R1/T1R3 Receptor.

The study aimed to explore umami peptides derived from protein hydrolysates of Morchella esculenta. According to the electronic tongue and sensory evaluation, the ultrafiltration fractions (<3 kDa) of the protein hydrolysates exhibited the strongest umami taste. The overall flavor of the screened fractions was significantly improved after the Maillard reaction, based on the electronic nose and electronic tongue analyses, and the content of total free amino acid increased from 387.35 to 589.30 µg/mL. A total of 37 peptides with high confidence were identified from the fractions using LC-MS/MS. Additionally, two novel umami peptides were screened through bioinformatics and molecular docking, and their recognition threshold was 0.43 (EYPPLGRFA) and 0.52 mmol/L (TVIDAPGHRDFI), respectively. In addition, molecular docking analysis revealed that the key binding sites, such as Ser148, Leu51, Arg327, and Leu468 in T1R1/T1R3 contributed to docking, and hydrogen bonding and hydrophobic interactions were the dominant interaction forces between the two umami peptides and T1R1/T1R3 receptor. This study contributes to the development and utilization of Morchella esculenta in flavored foods.

Comparison of chemical compositions, antioxidant activities, and acetylcholinesterase inhibitory activities between coffee flowers and leaves as potential novel foods.

This study aimed to compare chemical compositions, antioxidant activities, and acetylcholinesterase inhibitory activities of coffee flowers (ACF) and coffee leaves (ACL) with green coffee beans (ACGB) of Coffea Arabica L. The chemical compositions were determined by employing high-performance liquid chromatography-mass spectroscopy (HPLC-MS) and gas chromatography-mass spectroscopy (GC-MS) techniques. Antioxidant effects of the components were evaluated using DPPH and ABTS radical scavenging assays, and the ferric reducing antioxidant power (FRAP) assay. Their acetylcholinesterase inhibitory activities were also evaluated. The coffee sample extracts contained a total of 214 components identified by HPLC-MS and belonged to 12 classes (such as nucleotides and amino acids and their derivatives, tannins, flavonoids, alkaloids, benzene, phenylpropanoids, and lipids.), where phenylpropanoids were the dominant component (>30%). The contents of flavonoids, alkaloids, saccharides, and carboxylic acid and its derivatives in ACF and ACL varied significantly (p < .05) compared to similar components in ACGB. Meanwhile, 30 differentially changed chemical compositions (variable importance in projection [VIP] > 1, p < .01 and fold change [FC] > 4, or <0.25), that determine the difference in characteristics, were confirmed in the three coffee samples. Furthermore, among 25 volatile chemical components identified by GC-MS, caffeine, n-hexadecanoic acid, 2,2'-methylenebis[6-(1,1-dimethylethyl)-4-methyl-phenol], and quinic acid were common in these samples with caffeine being the highest in percentage. In addition, ACL showed the significantly highest (p < .05) DPPH radical scavenging capacity with IC50 value of 0.491 ± 0.148 mg/ml, and acetylcholinesterase inhibitory activity with inhibition ratio 25.18 ± 2.96%, whereas ACF showed the significantly highest (p < .05) ABTS radical scavenging activity with 36.413 ± 1.523 mmol trolox/g Ex. The results suggested that ACL and ACF had potential values as novel foods in the future.

Structural Characteristics, Rheological Properties, and Antioxidant and Anti-Glycosylation Activities of Pectin Polysaccharides from Arabica Coffee Husks.

As primary coffee by-products, Arabica coffee husks are largely discarded during coffee-drying, posing a serious environmental threat. However, coffee husks could be used as potential material for extracting pectin polysaccharides, with high bioactivities and excellent processing properties. Thus, the present study aimed to extract the pectin polysaccharide from Arabica coffee husk(s) (CHP). The CHP yield was calculated after vacuum freeze-drying, and its average molecular weight (Mw) was detected by gel permeation chromatography (GPC). The structural characteristics of CHP were determined by Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), proton nuclear magnetic resonance (1H NMR), and scanning electron microscopy (SEM). Additionally, the rheological and antioxidant properties of CHP and the inhibition capacities of advanced glycation end products (AGEs) with different concentrations were evaluated. The interaction mechanisms between galacturonic acid (GalA) and the AGE receptor were analyzed using molecular docking. The results demonstrated that the CHP yield was 19.13 ± 0.85%, and its Mw was 1.04 × 106 Da. The results of the structural characteristics results revealed that CHP was an amorphous and low-methoxyl pectic polysaccharide linked with an α-(1→6) glycosidic bond, and mainly composed of rhamnose (Rha, 2.55%), galacturonic acid (GalA, 45.01%), ß-N-acetyl glucosamine (GlcNAc, 5.17%), glucose (Glc, 32.29%), galactose (Gal, 6.80%), xylose (Xyl, 0.76%), and arabinose (Ara, 7.42%). The surface microstructure of CHP was rough with cracks, and its aqueous belonged to non-Newtonian fluid with a higher elastic modulus (G'). Furthermore, the results of the antioxidant properties indicated that CHP possessed vigorous antioxidant activities in a dose manner, and the inhibition capacities of AGEs reached their highest of 66.0 ± 0.35% at 1.5 mg/mL of CHP. The molecular docking prediction demonstrated that GalA had a good affinity toward AGE receptors by -6.20 kcal/mol of binding energy. Overall, the study results provide a theoretical basis for broadening the application of CHP in the food industry.

Isolation, identification and characterization of a novel antimicrobial peptide from Moringa oleifera seeds based on affinity adsorption.

This study aimed to characterize a novel antimicrobial peptide (AMP) obtained from Moringa oleifera seed protein hydrolysates. Cell membrane chromatography and live bacteria adsorption were combined into a single step to efficiently isolate the active fraction of the AMP. Five peptides were identified by LC-MS/MS, among which the MCNDCGA peptide (termed MOp3) showed the greatest inhibitory effect against Staphylococcus aureus [minimum inhibitory concentration (MIC): 2 mg/mL]. MOp3 was identified as a hydrophobic anionic AMP rich in ß-sheet structures with negligible hemolytic activity at 2.0 × MIC. MOp3 had good tolerance to salt solutions at 5 % and pH range 6.0-8.0, but was sensitive to high temperatures (>100 °C) and acid protease. Microscopic observation further revealed that MOp3 induced irreversible damage onto the cell membrane of S. aureus and interacted with dihydrofolate reductase and DNA gyrase by hydrogen bonding and hydrophobic interaction. These findings highlight the potential application of a new antimicrobial agent against S. aureus in the food industry.

Arginine-Containing Peptides Derived from Walnut Protein Against Cognitive and Memory Impairment in Scopolamine-Induced Zebrafish: Design, Release, and Neuroprotection.

The purpose of this study was to investigate the neuroprotective effect of Arg-containing peptides from walnut storage protein sequences in scopolamine-induced zebrafish and further to validate the potential neuroprotection of Arg-containing peptide enriched walnut hydrolysates prepared by in silico hydrolysis and controlled enzymatic release. Results showed that walnut derived Arg-containing peptides with high abundance and great bioactivity predicted by bioinformatics displayed potent neuroprotection in scopolamine-induced zebrafish, and regulation of neurotransmitter level and antioxidant enzyme activity might be the main target for Arg-containing peptides to exert neuroprotection. Notably, Arg-containing peptides (not free arginine) contributed greater neuroprotection, and the positive charge and cell-penetrating properties also affected their neuroprotection. Subsequently, Arg-containing peptides could be released efficiently from walnut protein following hydrolysis by trypsin, pepsin, papain, and thermolysin (bound arginine content: ranging from 110.43 ± 1.58 to 121.82 ± 1.02 mg/g). Among them, trypsin had excellent potential for releasing Arg-containing peptides in silico hydrolysis, and its hydrolysate was confirmed to have neuroprotective capacity, indicating that the combination of in silico hydrolysis and controlled enzymatic release might be an effective approach to obtain Arg-containing neuroprotective peptides.

Preparation, Sensory Characterization, and Umami-Enhancing Mechanism of Novel Peptide Glycoconjugates.

Previous studies supposed that Amadori rearrangement products (ARPs) of peptides might have better umami-enhancing abilities. To confirm this, five ARPs (EP-ARP, AH-ARP, EE-ARP, ß-AH-ARP, RFPHADF-ARP) were synthesized using a food-grade preparation method, and their chemical structures were clearly demonstrated by mass spectrometry and 1D/2D NMR. Sensory experiments showed that ARPs had better umami-enhancing abilities than the corresponding peptides in this research, though their enhancing performance varied. ARPs showed a synergistic effect with multiple umami substances (MSG and GMP), while their corresponding peptides did not. RFPHADF-ARP had good umami-enhancing capacity, despite that RFPHADF was a bitter peptide without any umami/umami-enhancing property. RFPHADF-ARP could bind to the T1R3, which is beneficial to the stability of the active conformation of the umami receptor. The introduction of glucose via the Maillard reaction increased the binding force of RFPHADF with the umami receptor by influencing the electron density distribution and offering more binding groups (hydroxide group).

Milk-clotting properties on bovine caseins of a novel cysteine peptidase from germinated Moringa oleifera seeds.

A cysteine peptidase was previously identified from germinated Moringa oleifera seeds, but its milk-clotting properties on bovine caseins was still unclear. In this study, this novel cysteine peptidase (MoCP) showed preferential activity on κ-casein (κ-CN), with greater hydrolytic activity compared with calf rennet, whereas weak hydrolysis of α-casein and ß-casein made MoCP suitable for application in cheesemaking and may yield various functional peptides. All 3 evaluated caseins were hydrolyzed to form relatively stable peptide bands within 3 h of proteolysis with MoCP. Cleavage sites were determined by gel electrophoresis, liquid chromatography mass spectrometry/mass spectrometry, and peptide sequencing, which revealed that cleavage of κ-CN by MoCP occurred at residue Ile129-Pro130 and generated a 14,895.37-Da peptide. The flocculation reaction between MoCP and κ-CN determined by 3-dimensional microscopy with super-depth of field revealed that the initial 30 min of reaction were key for milk coagulation, which may affect curd yield. Overall, the findings presented herein suggest that the cysteine peptidase from germinated M. oleifera seeds can be considered a promising plant-derived rennet alternative for use in cheese manufacture.

Protein function analysis of germinated Moringa oleifera seeds, and purification and characterization of their milk-clotting peptidase.

The present study aimed to investigate the biological functions of germinated M. oleifera seed proteins and to identify the identity of milk-clotting proteases. A total of 963 proteins were identified, and those with molecular weights between 10 and 30 kDa were most abundant. The identified proteins were mainly involved in energy-associated catalytic activity and metabolic processes, and carbohydrate and protein metabolisms. The numbers of proteins associated with the hydrolytic and catalytic activities were higher than the matured dry M. oleifera seeds reported previously. Of the identified proteins, proteases were mainly involved in the milk-clotting activity. Especially, a cysteine peptidase with a molecular mass of 17.727 kDa exhibiting hydrolase and peptidase activities was purified and identified. The identified cysteine peptidase was hydrophilic, and its secondary structure consisted of 27.60% alpha helix, 9.20% beta fold, and 63.20% irregular curl; its tertiary structure was also constructed using M. oleifera seed 2S protein as the protein template. The optimal pH and temperature of the purified protease were pH 4.0 and 60 °C, respectively. The protease had high acidic stability and good thermostability, thus could potentially be applied in the dairy industry.

Identification of novel autoantibodies for detection of malignant mesothelioma.

BACKGROUND: The malignant mesothelioma (MM) survival rate has been hampered by the lack of efficient and accurate early detection methods. The immune system may detect the early changes of tumor progression by responding with tumor-associated autoantibody production. Hence, in this study, we translated the humoral immune response to cancer proteins into a potential blood test for MM. METHODOLOGY/PRINCIPAL FINDINGS: A T7 phage MM cDNA library was constructed using MM tumor tissues and biopanned for tumor-associated antigens (TAAs) using pooled MM patient and normal serum samples. About 1008 individual phage TAA clones from the biopanned library were subjected to protein microarray construction and tested with 53 MM and 52 control serum samples as a training group. Nine candidate autoantibody markers were selected from the training group using Tclass system and logistic regression statistical analysis, which achieved 94.3% sensitivity and 90.4% specificity with an AUC value of 0.89 in receiver operating characteristic analysis. The classifier was further evaluated with 50 patient and 50 normal serum samples as an independent blind validation, and the sensitivity of 86.0% and the specificity of 86.0% were obtained with an AUC of 0.82. Sequencing and BLASTN analysis of the classifier revealed that five of these nine candidate markers were found to have strong homology to cancer related proteins (PDIA6, MEG3, SDCCAG3, IGHG3, IGHG1). CONCLUSIONS/SIGNIFICANCE: Our results indicated that using a panel of 9 autoantibody markers presented a promising accuracy for MM detection. Although the results need further validation in high-risk groups, they provided the potentials in developing a serum-based assay for MM diagnosis.