中药双向发酵液制备及其抗衰老、保湿、美白功效研究

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Process optimization and anti-aging, moisturizing and whitening effects of traditional Chinese medicine bi-directional fermentation broth

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doi:10.3969/j.issn.2097-2806.2023.05.005

Abstract:

In this study, the bi-directional fermentation process of Chinese Ganoderma lucidum, Dendrobium officinale, Polygonatum chinensis and Tuberose was optimized, and the optimal fermentation broth was prepared. Activity inhibition, Aquaporin3 （AQP3） content, 2, 2’-diazo-bis-3-ethylbenzothiazoline-6-sulfonic acid （ABTS） free radical scavenging rate, 1, 1-diphenyl-2-trinitrophenylhydrazine （DPPH） free radical scavenging rate and melanin content in mouse melanin B16-F10 cells were measured. The results of the study find that the bi-directional fermentation broth of traditional Chinese medicine can significantly inhibit protein non-enzymatic glycosylation, tyrosinase activity, and melanin content in mouse B16-F10 cells （P<0.05）. It can also significantly scavenge DPPH and ABTS free radicals, and upregulate AQP3 protein expression in keratinocytes （P<0.01）. Therefore, this paper concludes that the bi-directional fermentation broth of traditional Chinese medicine screened by optimizing the bi-directional fermentation process has a significant effect on inhibiting protein non-enzymatic glycosylation, tyrosinase activity and melanin content of mouse melanin B16-F10 cells, upregulating AQP3 protein expression, and scavenging ABTS free radicals and DPPH free radicals in vitro. It provides a new direction and basis for the development of cosmetic raw materials.

Key words: bi-directional fermentation, protein non-enzymatic glycosylation, tyrosinase, free radicals, aquaporin AQP3

CLC Number:

TQ658

Guo Fangyu, Han Tingting, Wang Xiaona, Chen Yurong, Wang Xiaomei, Yang Suzhen. Process optimization and anti-aging, moisturizing and whitening effects of traditional Chinese medicine bi-directional fermentation broth[J].China Surfactant Detergent & Cosmetics, 2023, 53(5): 523-531.

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样品编号	碳源（w/%）	氮源（w/%）	无机盐（w/%）	样品编号	碳源（w/%）	氮源（w/%）	无机盐（w/%）
1	1%葡萄糖	0.5%蛋白胨	0.5% NaCl，0.02% CaCO₃	4	1%蔗糖	0.5%酵母粉	0.5% NaCl，0.02% CaCO₃
2	1%葡萄糖	0.5%酵母粉		5	1%淀粉	0.5%蛋白胨
3	1%蔗糖	0.5%蛋白胨		6	1%淀粉	0.5%酵母粉

样品编号	w/%
样品编号	淀粉	酵母粉	NaCl	CaCO₃
1	1	0.5	0.3	0.02
2	1	1	0.5
3	1	1.5	0.7
4	2	0.5	0.5
5	2	1	0.7
6	2	1.5	0.3
7	3	0.5	0.7
8	3	1	0.3
9	3	1.5	0.5

样品编号	w/%
样品编号	铁皮石斛	黄精	晚香玉
1	1	0.1	0.05
2	1	0.2	0.1
3	1	0.4	0.2
4	2	0.1	0.1
5	2	0.2	0.2
6	2	0.4	0.05
7	3	0.1	0.2
8	3	0.2	0.05
9	3	0.4	0.1

序号	组别	信息		检测系统
序号	组别	样品名称	样品量	检测项目	检测模型	检测方法
组1	阴性对照组	SC		AQP3	Keratinocyte	IF
组1	样品组	中药双向发酵液	0.05%（w/%）	AQP3	Keratinocyte	IF
组2	阴性对照组	SC（1% DMSO）		AQP3	Keratinocyte	IF
组2	阳性对照组	维甲酸	2.5 μmol/L	AQP3	Keratinocyte	IF

pH 7.4缓冲溶液/mL

0.1%酪氨酸溶液/mL

0.3 g/mL酪氨酸酶溶液/mL

组别	样品信息		检测系统
组别	组别名称	样品量	检测项目	检测模型	检测方法
空白对照组	空白对照		黑素含量	B16-F10	吸光度法
阴性对照组	阴性对照（茶碱）	50 μg/mL
阳性对照组	阳性对照（α-熊果苷）	125 μg/mL
样品组	中药双向发酵液	2.5%，5%，10%

因素	淀粉含量（A）	酵母粉含量（B）	NaCl含量（C）	多糖质量浓度/（mg/mL）
1	1	1	1	0.35
2	1	2	2	0.68
3	1	3	3	0.82
4	2	1	2	0.40
5	2	2	3	0.77
6	2	3	1	0.91
7	3	1	3	0.31
8	3	2	1	0.60
9	3	3	2	0.71
K₁	0.650	0.353	0.687
K₂	0.760	0.683	0.663
K₃	0.607	0.980	0.667
极差	0.153	0.627	0.024

因素	铁皮石斛含量（A）	黄精含量（B）	晚香玉含量（C）	多糖质量浓度/（mg/mL）
1	1	1	1	5.11
2	1	2	2	5.45
3	1	3	3	5.23
4	2	1	2	5.83
5	2	2	3	5.98
6	2	3	1	5.79
7	3	1	3	5.38
8	3	2	1	5.55
9	3	3	2	5.21
K₁	5.263	5.440	5.483
K₂	5.867	5.660	5.497
K₃	5.380	5.410	5.530
极差	0.604	0.250	0.047

序号	组别	样品名称	样品量	平均荧光强度/a.u	SD	P值
组1	阴性对照	SC		7.12	0.46
组1	样品组	中药双向发酵液	0.05%（质量分数）	19.97	2.73	0.001
组2	阴性对照	SC（0.1% DMSO）		8.65	0.93
组2	阳性对照	维甲酸	2.5 μmol/L	14.95	1.62	0.004

组别名称	黑色素含量/mg	标准偏差	抑制率/%	标准偏差/%	P值
组别名称	黑色素含量/mg	标准偏差	抑制率/%	标准偏差/%	与NC比较
空白对照	3.32	0.06			0.008^##
阴性对照	3.63	0.06
2.5%中药双向发酵液	3.95	0.06	-8.80	1.74	0.006^**
5%中药双向发酵液	1.48	0.03	59.19	0.75	0.000^**
10%中药双向发酵液	1.33	0.09	63.37	2.54	0.000^**
α-熊果苷	2.77	0.10	23.85	2.68	0.000^**

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添加试剂	受试样品/mL	无水乙醇/mL	DPPH溶液/mL
样品组	1.0	0	1.0
阳性对照组	1.0	0	1.0
阴性对照组	0	1.0	1.0
空白对照组	1.0	1.0	0

发酵时间/d	多糖质量浓度/（mg/mL）
1	5.68
3	5.85
5	5.74
7	5.33