[논문]셀로비오스의 미백화장품 소재 가능성 평가를 위한 멜라닌 세포에서 자가포식 및 멜라닌 생성 효능 연구

차병선; 이석주; 정소영; 이소민; 이상훈; 빈범호; 곽병문; 허효진

doi:10.15230/scsk.2022.48.4.365

셀로비오스의 미백화장품 소재 가능성 평가를 위한 멜라닌 세포에서 자가포식 및 멜라닌 생성 효능 연구
Evaluation of the Potential of Cellobiose as a Material for Whitening Cosmetics based on Autophagy and Melanin Production Efficacy in Melanocytes 원문보기

大韓化粧品學會誌 = Journal of the society of cosmetic scientists of Korea, v.48 no.4, 2022년, pp.365 - 372

차병선 (아주대학교 생명과학과) , 이석주 (아주대학교 응용생명공학과) , (아주대학교 응용생명공학과) , 정소영 (아주대학교 응용생명공학과) , 이소민 (아주대학교 생명과학과) , (아주대학교 생명과학과) , 이상훈 (아주대학교 생명과학과) , (아주대학교 응용생명공학과) , 빈범호 (아주대학교 생명과학과) , 곽병문 (아주대학교 생명과학과) , 허효진 (아주대학교 응용생명공학과)

초록
AI-Helper

셀로비오스(cellobiose)는 식물 등에 널리 분포하는 셀롤로오스의 분해 과정에서 생성되는 이당류이다. 환원당인 셀로비오스는 2 개의 β-1,4 결합으로 결합한 구조를 가지며, 식물 등에 존재하는 천연물로서, 건강식품이나 식품의 감미료, 화장품 원료, 의약품 원료로서 활용되어지고 있다. 본 연구에서는 다양하게 유용될 셀로비오스를 기능성 화장품인 미백 화장품 소재로서의 가능성을 확인하고자, 자가포식 활성화 및 멜라닌 생성저해 효능을 검토하였다. 인간 유래 멜라닌 생성 세포로 알려진 MNT-1 세포에서 셀로비오스의 농도에 따른 독성 실험을 진행한 결과 20 mg/mL이하에서 세포독성이 없음을 확인하였고, 공초점 현미경을 통해 셀룰비오스 처리에 따른 자가포식 표지자인 microtubule-associated protein 1 light chain 3 (LC3) puncta의 증가가 확인되었다. 또한 LC3의 형태가 LC3-I에서 LC3-∏로 변환됨을 확인하였으며, 농도와 시간에 따라 LC3-II양이 증가하는 것을 확인하였다. 셀로비오스를 동일 세포에서 20 mg/mL 농도로 7 일간 처리하여 멜라닌 생성량을 분석한 결과, 50% 이상 멜라닌 생성 감소를 확인하였다. 추가적으로, 멜라닌 생성 관련 단백질TYR 및 TYRP1의 발현량을 western blot법을 이용하여 분석한 결과, 농도의존적으로 발현이 저해되어 멜라닌 생성이 감소함을 확인하였다. 이러한 연구 내용을 기반을 토대로 셀로비오스를 함유한 화장품 크림제형을 확보하여 4 주 동안 제형의 변화를 확인 해본 결과 성상이 고온에서 액채형태로 변화하였지만 pH는 변화하지 않았음을 확인하였다. 결론적으로, 자가포식 활성화 및 멜라닌 생성 억제 과정에 셀로비오스가 관여함을 확인하였으며, 이를 통해 미백 화장품 소재로서의 활용 가능성을 제안하고자 한다.

Abstract ▼ AI-Helper

Cellobiose is a dissacharide constituted by two glucose units joined by a β-('1,4') glycosidic bond that is produced by the decomposition of cellulose. This product exists naturally in plants and has been utilized in different industries as a food sweetener, and as a cosmetic and pharmaceutical material. In this study, the potential of cellobiose as a whitening cosmetic product was evaluated by analyzing autophagy induction and the inhibition of melanin production. A cytotoxicity test conducted in the human melanin-producing cell line MNT-1 with increasing concentrations of cellobiose revealed that this compound did not cause cytotoxicity at 20 mg/mL or less. Based on this, autophagy was firstly evaluated by immunostaining with the autophagy marker microtubule-associated protein 1 light chain 3 (LC3) after treatment with 20 mg/mL of cellobiose. The subsequent confocal microscopy analysis revealed an increase in LC3 puncta, indicating induction of autophagy. In addition, autophagy was further confirmed by western blot analysis, which demonstrated that cellobiose converted LC3-I to LC3- ∏ in a concentration- and time-dependent manners. An analysis of melanin contents after cellobiose treatment at a concentration of 20 mg/mL during 7 days revealed that melanin production was reduced by more than 50%. Additionally, the expression levels of melanogenesis-related proteins TYR and TYRP1 were markedly decreased after cellobiose treatment. Based on these studies, a cosmetic cream formulation containing cellobiose was prepared and the change in formulation was tested for 4 weeks, and it was confirmed that the appearance changed to liquid form at high temperature, but the pH did not change. In conclusion, the present research demonstrated that cellobiose activates autophagy and inhibits melanin production, and showed the potential of this product as a material for whitening cosmetics.

주제어

표/그림 (5)

표 Table 1. Moisturizing Cream Containing Cellobiose
그림 Figure 1. Effect of cellobiose treatment in cell viability and autophagosome formation in MNT-1 cells. (A) Cell viability of MNT-1 cells after treatment with increasing concentrations of cellobiose. (B) LC3 punta observed by confocal microscopy after treatment of MNT-1 cells with 20 mg/mL of glucose or cellobiose and immunostaining with LC3 antibody. Nuclei were stained with DAPI. The results are presented as the mean ± SD of the experiment (N = 3), ^*p < 0.05, ^**p < 0.01, ^***p < 0.005, compared to control.
그림 Figure 2. Cellobiose treatment led to increased autophagy in MNT-1 cells. (A) Results of the protein expression levels of LC3-I and LC3-Ⅱ in MNT-1 cells after treatment with increasing cellobiose concentrations. (B) Comparison of LC3-I and LC3-Ⅱ protein expression levels relative to Figure 2A after quantification with Image J software. (C) Results of the protein expression levels of LC3-I and LC3-Ⅱ in MNT-1 cells overtime after treatment with 20 mg/mL of cellobiose. (D) Comparison of LC3-I and LC3-Ⅱ protein expression levels relative to Figure 2C after quantification with Image J software. CTL, also known as control, was not treated with cellobiose. The results are presented as t he m ean ± SD of t he e xperiment (N = 3), ^*p < 0.05, ^**p < 0.01, ^***p < 0.005, compared to control.
그림 Figure 3. Cellobiose reduced melanin production and downregulated the expression of melanogenesis-associated proteins in MNT-1 cells. (A) Optical comparison of the melanin contents of MNT-1 cells after treatment with 20 mg/mL of cellobiose. (B) Quantification of the melanin contents relative to Figure 3A by melanin assay analysis. (C) Expression levels of melanogenesis-associated proteins TYR and TYRP1 in MNT-1 cells after treatment with 20 mg/mL of cellobiose. (D) Comparison of TYR and TYRP1 protein expression levels relative to Figure 3C after quantification with Image J software. CTL, also known as control, was not treated with cellobiose. The results are presented as the mean ± SD of the experiment (N = 3), ^*p < 0.05, ^**p < 0.01, ^***p < 0.005, compared to control.
그림 Figure 4. Stability of a cosmetic cream formulation containing cellobiose. The stability of creams formulated with 2% cellobiose was tested by monitoring the pH and the visible features of the products over 4 weeks.

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