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Author: search.filters.author.Sangtanoo, PapassaraSubject: search.filters.subject.Protein hydrolysate

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    Bee pollen peptides as potent tyrosinase inhibitors with anti-melanogenesis effects in murine b16f10 melanoma cells and zebrafish embryos
    (2024-12-01)
    Sangtanoo, Papassara
    ;
    Srimongkol, Piroonporn
    ;
    Saisavoey, Tanatorn
    ;
    Puthong, Songchan
    ;
    Buakeaw, Anumart
    One important functional food ingredient today, valued for its health properties and ability to prevent disease, is bee pollen, which comprises a combination of nectar, pollen from plants, and the secretions of bees. In this research, the tyrosinase (TYR) inhibiting abilities of the peptides derived from bee pollen protein hydrolysates are investigated. Various proteases were utilized to generate these peptides, followed by testing at different concentrations. Tyrosinase inhibition activity was detected in all cases, while the hydrolysate drawn from 5.0% w/v neutrase exhibited the best IC<inf>50</inf> value and was thus investigated further via ultrafiltration to separate the active fractions. The highest potential for tyrosinase inhibition was recorded for the fractions below 0.65 kDa. Subsequent purification steps via SEC and RP-HPLC led to the identification of the VDGYPAAGY (named VY-9) peptide via LC-Q-TOF-MS/MS in fraction F<inf>1–2</inf>, known for its non-toxic and hydrophobic characteristics albeit poor water solubility. The synthesized VY-9 peptide demonstrated competitive inhibition, with IC<inf>50</inf> values of 0.55 ± 0.03 µM for mono-phenolase and 2.54 ± 0.06 µM for di-phenolase activities, as confirmed by molecular docking analysis revealing dominant hydrogen bond interactions with TYR. Effective concentrations of 0.2–1.6 µM of VY-9 showed negligible cytotoxicity in B16F10 cells. Melanin synthesis suppression was examined via qRT-PCR, and western blot in MITF, TYR, TRP-1, and TRP-2. Cell death in zebrafish embryos was evaluated in vivo using a toxicity assay which revealed no significant influence from VY-9, while anti-melanogenic effects were observed when the concentration was 4 µM, suggesting bee pollen-derived peptides’ potential in cosmetic and pharmaceutical depigmentation applications.
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    Optimization, isolation, identification and molecular mechanisms in B16F10 melanoma cells of a novel tyrosinase inhibitory peptide derived from split gill mushrooms
    (2024-10-01)
    Arnamwong, Sarinya
    ;
    Kuptawach, Kittisak
    ;
    Sangtanoo, Papassara
    ;
    Srimongkol, Piroonporn
    ;
    Saisavoey, Tanatorn
    Hyperpigmentation often arises from an imbalance in melanogenesis, primarily due to the overexpression of tyrosinase (TYR). While the inhibition of TYR presents a common approach to skin whitening, it can lead to undesirable side effects. Thus, there is growing interest in safe and natural alternatives for TYR inhibition. Bioactive compounds and peptides sourced from split gill mushrooms hold promise in this regard. This study aims to optimize the conditions for papain-mediated hydrolysis of split gill mushroom protein to inhibit TYR activity, utilizing response surface methodology (RSM) and central composite design (CCD). Optimal conditions were determined at a temperature of 46.70 °C, a hydrolysis time of 217.09 min, and an enzyme-to-substrate ratio (E/S) of 1.1%. Under these conditions, the resulting hydrolysates exhibited significant TYR inhibition, with an IC<inf>50</inf> value of 117.86 μg/mL and a degree of hydrolysis (DH) of 87.97%. Further purification via ultrafiltration and RP-HPLC yielded a peptide, Tyr-Ala-Ser-Ile-Leu-Leu (YASILL or YL-6), identified through LC-Q-TOF-MS/MS, which competitively inhibited TYR. YL-6 demonstrated an IC<inf>50</inf> value of 3.97 mM for mono-phenolase activity and 6.75 mM for di-phenolase activity. Molecular docking analysis revealed hydrogen bonds and hydrophobic interactions between TYR and YL-6. Treatment of B16F10 cells with YL-6 across concentrations ranging from 10-3000 μM showed no cytotoxic effects.The inhibition of melanin synthesis was investigated via qRT-PCR along with Western blot in MITF, TYR, TRP-1, and TRP-2. The results obtained in this research may prove significant in guiding the development of commercially viable cosmetic products to whiten the skin.
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    Free radical scavenging properties and induction of apoptotic effects of fa fraction obtained after proteolysis of bioactive peptides from microalgae synechococcus sp. VDW
    (2019-01-01)
    Suttisuwan, Rutairat
    ;
    Phunpruch, Saranya
    ;
    Saisavoey, Tanatorn
    ;
    Sangtanoo, Papassara
    ;
    Thongchul, Nuttha
    This study aims to determine the antioxidant activity of bioactive peptides derived from Synechococcus sp. VDW cells cultured for 21 days. Synechococcus sp. VDW protein hydrolysates were prepared with trypsin and purified by ultrafiltration with molecular mass cut-off membranes of 10, 5 and 3 kDa. The M<3 kDa (FA) fraction had the highest 2,2'-azino- bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 2,2'-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities, with IC50 values of (11.5±0.3) and (13.6±0.2) μg/ mL, respectively. The FA fraction was separated by reversed phase HPLC to yield four subfractions (F1-4). The F4 subfraction showed the highest maximum ABTS radical scavenging activity (3.55±0.61) % and it was selected for further analysis by electrospray ionisation quadrupole time-of-flight mass spectrometry (ESI-Q-TOF-MS/MS) based on de novo peptide sequencing. Five antioxidant peptides were identified, of which AILESYSAGKTK had the highest ABTS radical scavenging activity. Furthermore, the FA fraction showed high cytotoxic activities against human cancer-derived cell lines, especially the colon cancer cell line (SW620) with an IC50 value of (106.6±21.5) μg/mL, but not the untransformed Wi38 cell line. The FA fraction activated the apoptotic pathway in SW620 cells after treatment for 24, 48 and 72 h, with the highest activities of caspases-3, -8 and -9 being observed after treatment for 72 h. These findings suggested that microalgae Synechococcus sp. VDW may be used to develop natural anticancer drugs.