多菌种联用发酵燕麦麸皮工艺优化及发用功效评价

doi:10.13560/j.cnki.biotech.bull.1985.2023-0163

摘要/Abstract

摘要：

为开发燕麦麸皮的综合利用价值，本研究采用单因素和正交试验法，优化了多菌种联用发酵法共提取燕麦麸皮β-葡聚糖和燕麦多肽的工艺条件，并对发酵提取液的抗氧化能力、抑菌性和应用于发用化妆品的效果进行评价。通过对发酵过程中枯草芽孢杆菌（Bacillus subtilis）与地衣芽孢杆菌（Bacillus licheniformis）活菌数的测定，确定了适合燕麦麸皮发酵的最优发酵组合为枯草芽孢杆菌、地衣芽孢杆菌与酵母菌（Saccharomyces cerevisiae）联用，接种比7∶3∶5。以多肽与β-葡聚糖综合得率为评定指标，通过单因素试验与四因素三水平正交试验，确定最佳发酵工艺条件为料液比1∶7.5、温度30℃、接种量2%、发酵时间42 h，β-葡聚糖和燕麦多肽得率分别为5.98%和16.21%。燕麦麸皮发酵液的DPPH、ABTS和羟自由基清除率分别为89.00%、99.50%和93.01%，IC₅₀分别为89.35、1.315和91.26 mg/mL。此外，发酵液对大肠埃希菌（Escherichia coli）、金黄色葡萄链球菌（Staphylostreptococcus aureus）和马拉色菌（Malassezia furfur）具有抑制效果，最小抑菌浓度分别为12.5、25和12.5 mg/mL。发酵提取液在增加发丝光泽度、修护毛鳞片、改善干梳理性、增加α-角蛋白还原能力方面均有显著效果。综合3种提取工艺的目标活性物质含量的检测及功效评价，实验发现发酵提取工艺相较于热提取法与酶解提取法，对β-葡聚糖与多肽的综合提取效率提升，且在抗氧化、抑菌、修护毛鳞片与还原角蛋白功能上有更高的活性，为燕麦麸皮在发用化妆品领域的应用提供依据。

关键词: 发酵, 燕麦麸皮, 多菌种, 工艺优化, 抗氧化, 抑菌, 发用功效评价

Abstract:

The fermentation conditions for co-extraction of β-glucan and polypeptide from oat bran by mixed bacteria and yeast culture were optimized using single factor and orthogonal test. The fermentation broth was analyzed for antioxidant and bacteriostatic performance, as well as application in hair protection. The optimal strain mixture for fermentation contained Bacillus subtilis, Bacillus licheniformis and Saccharomyces cerevisiae at an inoculation ratio of 7∶3∶5 by detecting of the viable number of B. subtilis and B. licheniformis during fermentation. The optimized fermentation conditions, resulting in β-glucan yield at 5.98% and oat polypeptide yield at 16.21%, were found to be the ratio of solid to liquid at 1∶7.5, temperature at 30℃, inoculation volume at 2% and fermentation time for 42 h and the main and secondary order of the effects of various factors on the fermentation yield of oat bran is material liquid ratio > inoculation amount > fermentation time > fermentation temperature. The fermentation broth reduced the radical formation as the radical scavenging rate of DPPH, ABTS and hydroxyl radical was 89.00%, 99.50% and 93.01%, respectively, with IC₅₀ of 89.35, 1.315 and 91.26 mg/mL, respectively. The antibacterial activity of the fermentation broth against Escherichia coli, Staphylostreptococcus aureus and Malassezia furfur was analyzed and the minimum inhibitory concentration was 12.5, 25 and 12.5 mg/mL, respectively. The fermentation broth had significant effect on increasing hair glossiness, repairing hair scales, reducing dry hair combing force and improving reducibility of α-keratin. Based on the detection of the target active substance content and the evaluation of its efficacy in three extraction processes, the experimental results showed that the fermentation extraction process improved the comprehensive extraction efficiency of β-glucan and peptides compared to the heating extraction method and enzymatic extraction method, and had higher activity in antioxidant, antibacterial, repairing hair scales, and reducing keratin functions.

Key words: fermentation, oat bran, multiple strains, process optimization, antioxidation, bacteriostasis, hair efficacy evaluation

张岳一, 兰社益, 裴海闰, 封棣. 多菌种联用发酵燕麦麸皮工艺优化及发用功效评价[J]. 生物技术通报, 2023, 39(9): 58-70.

ZHANG Yue-yi, LAN She-yi, PEI Hai-run, FENG Di. Process Optimization of Multi-strain Fermented Oat Bran and Hair Efficacy Evaluation[J]. Biotechnology Bulletin, 2023, 39(9): 58-70.

图/表 13

表1 正交试验设计

Table 1 Orthogonal experimental design

水平 Level	因素 Factor
水平 Level	A料液比 Solid-liquid ratio	B温度 Temperature/℃	C时间 Time/h	D接种量 Inoculation amount/%
1	1∶5	28	30	2
2	1∶7.5	30	36	3
3	1∶10	32	42	4

图1 枯草芽孢杆菌与地衣芽孢杆菌在接种比9∶1（A）、7∶3（B）、5∶5（C）、3∶7（D）和1∶9（E）时的生长曲线

Fig. 1 Growth curves of Bacillus subtilis and Bacillus licheniformis in inoculation proportion of 9∶1（A）, 7∶3（B）,5∶5（C）, 3∶7（D）and 1∶9（E）

图2 发酵时间、接种量、温度和料液比对多肽、β-葡聚糖得率的影响

Fig. 2 Effects of fermentation time, inoculation volume, temperature and solid-liquid ratio on peptides and β-glucan yield

表2 正交试验结果及分析

Table 2 Orthogonal test results and analysis

试验号 Test number	因素 Factor				β-葡聚糖得率 β-glucan yield /%	多肽得率 Peptide yield /%	综合得率 Comprehensive yield/%
试验号 Test number	A料液比 Solid-liquid ratio	B温度 Temperature/℃	C时间Time/h	D接种量 Inoculation amount/%	β-葡聚糖得率 β-glucan yield /%	多肽得率 Peptide yield /%	综合得率 Comprehensive yield/%
1	1	1	1	1	2.012	9.519	5.765
2	1	2	2	2	3.960	11.675	7.817
3	1	3	3	3	4.535	12.217	8.376
4	2	1	2	3	5.525	14.003	9.764
5	2	2	3	1	6.055	14.312	10.183
6	2	3	1	2	5.595	15.534	10.564
7	3	1	3	2	5.825	15.621	10.723
8	3	2	1	3	5.502	13.219	9.360
9	3	3	2	1	2.450	11.322	6.886
K1	7.319	8.750	8.563	7.611
K2	10.170	9.120	8.155	9.701
K3	8.989	8.608	9.760	9.166
R	2.851	0.412	1.605	2.090

图3 不同提取方法多肽、β-葡聚糖得率对比

Fig. 3 Comparison of peptide and β-qlucan yields by diffe-rent extraction processes

图4 不同提取方法DPPH清除率与IC50对比 HE：葡聚糖提取液；EE：多肽提取液；FE：燕麦麸皮发酵液。* P<0.05；** P <0.01；*** P <0.001。下同

Fig. 4 Comparison of DPPH clearance rate and IC50 under different extraction methods HE: Glucan extract; EE: peptide extract; FE: oat bran fermentation extract. * P <0.05; ** P <0.01; *** P <0.001. The same below

图5 不同提取方法ABTS清除率与 IC50 对比

Fig. 5 Comparison of ABTS clearance rate and IC50 under different extraction methods

图6 不同提取方法的羟自由基清除率与 IC50 对比

Fig. 6 Comparison of hydroxy radical scavenge rates and IC50 under different extraction methods

表3 不同提取液抑菌直径与最小抑菌浓度

Table 3 Bacteriostatic diameter and minimum inhibitory concentration of different extract

组别 Group	抑菌种类 Strains	抑菌圈直径 Antibacterial circle diameter/mm	最小抑菌浓度（MIC） Minimum inhibitory concentration/（mg·mL^-1）
对照组 Control group	金黄色葡萄链球菌	6.00	-
	大肠杆菌	6.00	-
	马拉色菌	6.00	-
葡聚糖提取液 Heating extract（HE）	金黄色葡萄链球菌	6.00	-
	大肠杆菌	6.00	-
	马拉色菌	6.00	-
多肽提取液 Enzymatic extract（EE）	金黄色葡萄链球菌	11.19±0.37	100
	大肠杆菌	13.92±0.26	100
	马拉色菌	9.69±0.26	50
燕麦麸皮发酵液 Fermentation extract（FE）	金黄色葡萄链球菌	15.05±0.12	25
	大肠杆菌	16.05±0.27	12.5
	马拉色菌	13.60±0.37	12.5

图7 不同提取液浸渍发丝光泽度变化

Fig. 7 Changes in the glossiness of hair soaked with different extract

图8 不同提取液浸渍发丝干梳理功（A）与干梳理最大负荷（B）

Fig. 8 Dry carding work（A）and maximum load（B）of dry carding for hair soaked with different extract

图9 不同提取液浸渍发丝相对螺旋含量

Fig. 9 Relative helix content of hair soaked with different extract

图10 经蒸馏水（A）、葡聚糖提取液（B）、多肽提取液（C）和发酵液（D）浸提发丝的扫描电镜图

Fig. 10 SEM images of hairs soaked with distilled water（A）, glucan extract（B）, peptide extract（C）and fermentation extract（D）

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