混菌发酵白酒糟生产含功能成分饲料发酵条件的优化

doi:10.13560/j.cnki.biotech.bull.1985.2021-0153

摘要/Abstract

摘要：

以蒸汽爆破预处理酒糟为主要原料,前期筛选菌株作为发酵菌种,以期优化发酵条件并生产富含功能成分酒糟饲料。将粗蛋白含量和烟酸含量作为主要指标,粗纤维含量和乙偶姻含量作为次要指标,通过单因素试验与正交试验结合探究麸皮添加量、装料量、初始pH、发酵时间、发酵温度及接种量对混菌固态发酵白酒糟的影响,进而优化酒糟发酵条件。结果表明酒糟发酵最佳工艺条件为:基料中麸皮添加量5%、装料量80 g、初始pH 3.70、发酵时间5 d、发酵温度30℃、接种量10%。与优化前相比,粗蛋白含量提高了1.56%（35.21%）,粗纤维含量降低了33.11%（12.73%）,烟酸含量增长率达到290.32%（1.21 mg/g酒糟）,乙偶姻含量保持基本不变（37.30 mg/g酒糟）。

关键词: 混菌发酵, 白酒糟, 饲料, 功能成分, 优化

Abstract:

In order to optimize the fermentation conditions and produce distiller’s grains rich in functional components,the steam explosion pretreatment of distiller’s grains was used as the main raw material,and the strains screened in the early stage was as fermentation strains. The effects of bran addition amount,loading amount,initial pH value,fermentation time,fermentation temperature,and inoculum amount,on the solid-state fermentation of distiller’s grains were studied by taking the crude protein content and nicotinic acid content as the main indexes,and the crude fiber content and acetoin content as the secondary indexes. The results showed that the optimal fermentation conditions were as follows:bran addition 5%,loading 80 g,initial pH 3.70,fermentation time 5 d,fermentation temperature 30℃ and inoculum 10%. Compared with those before optimization,the content of crude protein increased by 1.56%（35.21%）,the content of crude fiber decreased by 33.11%（12.73%）,the content of nicotinic acid increased by 290.32%（1.21 mg/g distiller’s grains）,and the content of acetoin remained unchanged（37.30 mg/g distiller’s grains）.

Key words: mixed fermentation, distiller’s grains, feed, functional component, optimization

范恩帝, 蒋梦迎, 冯敏雪, 陈叶福, 肖冬光, 郭学武. 混菌发酵白酒糟生产含功能成分饲料发酵条件的优化[J]. 生物技术通报, 2021, 37(12): 91-103.

FAN En-di, JIANG Meng-ying, FENG Min-xue, CHEN Ye-fu, XIAO Dong-guang, GUO Xue-wu. Optimization of Fermentation Conditions for Production of Feed Containing Functional Components from Distiller’s Grains by Mixed Bacteria Fermentation[J]. Biotechnology Bulletin, 2021, 37(12): 91-103.

图/表 12

参考文献 38

[1]	Savvidou SE, Beal JD, Brooks PH, et al. Liquid feed fermented with Lactobacillus salivarius reduces susceptibility of broiler chickens to Salmonella enterica typhimurium Sal 1344 nalr[J]. Proc BrSoc Anim Sci, 2009, 2009:201.
[2]	王晓力, 孙尚琛, 王永刚, 等. 饲料原料白酒糟基本成分测定及评价[J]. 粮油加工:电子版, 2015(5):62-65.
	Wang XL, Sun SC, Wang YG, et al. Analysis and evaluation of basic elements of distiller’s grains for fodder[J]. Cereals Oils Process:Electron Version, 2015(5):62-65.
[3]	张丽华, 王小媛, 李昌文, 等. 酒糟再利用的研究进展[J]. 食品与发酵工业, 2017, 43(11):250-256.
	Zhang LH, Wang XY, Li CW, et al. Recent advances in comprehensive utilization of grain stillage[J]. Food Ferment Ind, 2017, 43(11):250-256.
[4]	张轩, 赵述淼, 陈海燕, 等. 酿酒酵母固态发酵白酒糟生产蛋白饲料的研究[J]. 饲料工业, 2012, 33(19):27-31.
	Zhang X, Zhao SM, Chen HY, et al. Study on producing protein feed through the solid state fermentation of distiller’s grains with Saccharomyces cerevisiae[J]. Feed Ind, 2012, 33(19):27-31.
[5]	王晓力. 白酒糟生产高蛋白饲料研究进展及前景[J]. 中兽医医药杂志, 2013, 32(6):34-36.
	Wang XL. Research progress and prospects of production of protein feed by using distiller’s grains[J]. J Tradit Chin Vet Med, 2013, 32(6):34-36.
[6]	杨晓慧, 李桂明, 林树乾, 等. 酒糟作为饲料在家禽生产中的研究进展[J]. 家禽科学, 2012(10):10-13.
	Yang XH, Li GM, Lin SQ, et al. Research progress of distiller’s grains as feed in poultry production[J]. Poult Sci, 2012(10):10-13.
[7]	谭之磊, 刘建辉, 马凯, 等. 白酒糟综合利用新技术[J]. 广东化工, 2015, 42(7):72-73.
	Tan ZL, Liu JH, Ma K, et al. New technologies for comprehensive utilization of distillers grains[J]. Guangdong Chem Ind, 2015, 42(7):72-73.
[8]	吴正肖. 复合微生物肥料研制与效果研究[D]. 贵阳:贵州大学, 2017.
	Wu ZX. Development and evaluation of compound microbial fertilizers[D]. Guiyang:Guizhou University, 2017.
[9]	钱晓飞, 汤家铭. 特殊配方饲料引起实验动物体重增长缓慢的原因探讨[J]. 中国比较医学杂志, 2007, 17(12):752-754.
	Qian XF, Tang JM. Effect of peculiar formula feed on the body weight of laboratory animals[J]. Chin J Comp Med, 2007, 17(12):752-754.
[10]	齐志国, 郭江鹏, 陈余, 等. 功能性饲料及其功能因子概述[J]. 饲料研究, 2017(12):1-7, 12.
	Qi ZG, Guo JP, Chen Y, et al. Summary of functional feed and its functional factors[J]. Feed Res, 2017(12):1-7, 12.
[11]	Kemppainen K, Rommi K, Holopainen U, et al. Steam explosion of Brewer’s spent grain improves enzymatic digestibility of carbohydrates and affects solubility and stability of proteins[J]. Appl Biochem Biotechnol, 2016, 180(1):94-108. doi: 10.1007/s12010-016-2085-9 pmid: 27085356
[12]	闭帅珠, 彭林才, 陈克利. 蔗渣纤维素乙醇的预处理技术研究进展[J]. 生物质化学工程, 2016, 50(2):53-60.
	Bi SZ, Peng LC, Chen KL. Advances in pretreatment technology of sugarcane bagasse for bioethanol production[J]. Biomass Chem Eng, 2016, 50(2):53-60.
[13]	Katsimpouras C, Dimarogona M, Petropoulos P, et al. A thermostable GH26 endo-β-mannanase from Myceliophthora thermophila capable of enhancing lignocellulose degradation[J]. Appl Microbiol Biotechnol, 2016, 100(19):8385-8397. doi: 10.1007/s00253-016-7609-2 URL
[14]	Pachauri P, Chakradhari Y, Veeramani A. Study of activators and inhibitors on cellulase production from isolated Trichoderma koningii for effective saccharification of sugarcane bagasse[J]. Biofuels, 2020, 11(6):733-739. doi: 10.1080/17597269.2017.1400856 URL
[15]	Reddy G, Golla N, Dileep Kumar K, et al. Cellulase production by Aspergillus Niger on different natural lignocellulosic substrates[J]. International Journal of Current Microbiology and Applied Sciences, 2015, 4:835-845.
[16]	Yang F, Gong YF, Liu G, et al. Enhancing cellulase production in thermophilic fungus Myceliophthora thermophila ATCC42464 by RNA interference of cre1 gene expression[J]. J Microbiol Biotechnol, 2015, 25(7):1101-1107. doi: 10.4014/jmb.1501.01049 URL
[17]	Seo J, Park E, Leek SS, et al. Isolation of endo-1, 4-b-D-glucanase producing Bacillus subtilis sp. from fermented foods and enhanced enzyme production bydeveloping the mutant strain[J]. Indian J Animal Res, 2017, 51(4):785-790.
[18]	Ploss TN, Reilman E, Monteferrante CG, et al. Homogeneity and heterogeneity in amylase production by Bacillus subtilis under different growth conditions[J]. Microb Cell Fact, 2016, 15:57. doi: 10.1186/s12934-016-0455-1 pmid: 27026185
[19]	Guo X, Chen DD, Peng KS, et al. Identification and characterization of Bacillus subtilis from grass carp(Ctenopharynodon idellus)for use as probiotic additives in aquatic feed[J]. Fish Shellfish Immunol, 2016, 52:74-84. doi: 10.1016/j.fsi.2016.03.017 URL
[20]	Dai JY, Cheng L, He QF, et al. High acetoin production by a newly isolated marine Bacillus subtilis strain with low requirement of oxygen supply[J]. Process Biochem, 2015, 50(11):1730-1734. doi: 10.1016/j.procbio.2015.07.010 URL
[21]	Jiang RR, Zhao GP, Chen JL, et al. Effect of dietary supplemental nicotinic acid on growth performance, carcass characteristics and meat quality in three genotypes of chicken[J]. J Anim Physiol Anim Nutr:Berl, 2011, 95(2):137-145.
[22]	Yang ZQ, Bao LB, Zhao XH, et al. Nicotinic acid supplementation in diet favored intramuscular fat deposition and lipid metabolism in finishing steers[J]. Exp Biol Med:Maywood, 2016, 241(11):1195-1201. doi: 10.1177/1535370216639395 URL
[23]	Kurcz A, Błażejak S, Kot AM, et al. Application of industrial wastes for the production of microbial single-cell protein by fodder yeast Candida utilis[J]. Waste Biomass Valorization, 2018, 9(1):57-64. doi: 10.1007/s12649-016-9782-z URL
[24]	Plumed-Ferrer C, von Wright A. Antimicrobial activity of weak acids in liquid feed fermentations, and its effects on yeasts and lactic acid bacteria[J]. J Sci Food Agric, 2011, 91(6):1032-1040. doi: 10.1002/jsfa.4278 URL
[25]	Yang F, Hou C, Zeng X, et al. The use of lactic Acid bacteria as a probiotic in Swine diets[J]. Pathogens, 2015, 4(1):34-45. doi: 10.3390/pathogens4010034 URL
[26]	刘露, 张雁, 魏振承, 等. 肠道益生菌体外发酵山药低聚糖产短链脂肪酸的研究[J]. 食品科学技术学报, 2019, 37(4):49-56.
	Liu L, Zhang Y, Wei ZC, et al. Study on production of short chain fatty acids from yam oligosaccharides by intestinal probiotics fermentation in vitro[J]. J Food Sci Technol, 2019, 37(4):49-56.
[27]	李旭业, 董扬, 尤海洋, 等. 不同感官分级方法对全株玉米青贮饲料评价比较[J]. 现代畜牧科技, 2017(3):1-3.
	Li XY, Dong Y, You HY, et al. Evaluation and comparison of different sensory grading methods for whole plant corn silage[J]. Mod Animal Husb Sci Technol, 2017(3):1-3.
[28]	李芳香, 张稳, 郁建平, 等. 茅台酱香型酒糟基本成分的测定与分析[J]. 贵州农业科学, 2016, 44(9):114-116.
	Li FX, Zhang W, Yu JP, et al. Determination and analysis of basic components of vinasse with Moutai-flavor type[J]. Guizhou Agric Sci, 2016, 44(9):114-116.
[29]	中华人民共和国国家质量监督检验检疫总局, 中国国家标准化管理委员会. 中华人民共和国推荐性国家标准:饲料中粗纤维的含量测定过滤法 GB/T 6434—2006[S]. 北京: 中国标准出版社, 2006.
	General Administration of Quality Supervision, Inspection and Quarantine of the People’s Republic of China, Standardization Administration of the People’s Republic of China. National Standard（Recommended）of the People’s Republic of China:Feeding stuffs-Determination of crude fiber content Method with intermediate filtration. GB/T 6434—2006[S]. Beijing: Standards Press of China, 2006.
[30]	李金林, 许迟, 何立荣, 等. 近红外光谱分析技术在饲料检测领域的应用研究[J]. 饲料研究, 2017(5):10-12.
	Li JL, Xu C, He LR, et al. Application of near infrared spectroscopy in feed detection[J]. Feed Res, 2017(5):10-12.
[31]	中华人民共和国国家质量监督检验检疫总局, 中国国家标准化管理委员会. 中华人民共和国国家标准:饲料添加剂烟酸 GB 7300—2017[S]. 北京: 中国标准出版社, 2017.
	General Administration of Quality Supervision, Inspection and Quarantine of the People’s Republic of China, Standardization Administration of the People’s Republic of China. National Standard（Mandatory）of the People’s Republic of China:Feed additive—Nicotinic acid. GB 7300—2017[S]. Beijing: Standards Press of China, 2017.
[32]	Chen JC, Chen QH, Guo Q, et al. Simultaneous determination of acetoin and tetramethylpyrazine in traditional vinegars by HPLC method[J]. Food Chem, 2010, 122(4):1247-1252. doi: 10.1016/j.foodchem.2010.03.072 URL
[33]	孙东伟, 刘军, 牛广杰. 多菌种固态发酵酒糟生产菌体饲料蛋白的研究[J]. 中国饲料, 2010(7):38-39, 43.
	Sun DW, Liu J, Niu GJ. Study on the condition of disstiller’s grains fermentation with mixed strains[J]. China Feed, 2010(7):38-39, 43.
[34]	张连忠. 酵母固态发酵过程中营养物质变化的研究[J]. 饲料工业, 2011, 32(2):44-46.
	Zhang LZ. Studies on changes of nutrients in solid state fermentation of yeast[J]. Feed Ind, 2011, 32(2):44-46.
[35]	张谦, 贾佳, 林智, 等. 产脂肪酶黑曲霉摇瓶发酵条件优化研究[J]. 生物技术通报, 2015, 31(12):227-233. doi: 10.13560/j.cnki.biotech.bull.1985.2015.12.033
	Zhang Q, Jia J, Lin Z, et al. The optimization of flask fermentation conditions for the production of extracellular lipase from Aspergillus niger[J]. Biotechnol Bull, 2015, 31(12):227-233. doi: 10.13560/j.cnki.biotech.bull.1985.2015.12.033
[36]	张建华. 酒糟发酵蛋白质饲料菌种筛选的研究[J]. 粮食与饲料工业, 2010(8):46-48.
	Zhang JH. Studies on selection of bacterium for protein feed fermented from distiller’s grain[J]. Cereal Feed Ind, 2010(8):46-48.
[37]	郭素环, 周碧君, 文明, 等. 白酒糟发酵菌种组合的筛选[J]. 饲料工业, 2012, 33(15):17-21.
	Guo SH, Zhou BJ, Wen M, et al. Screening of fermentation strains of distiller’s grains[J]. Feed Ind, 2012, 33(15):17-21.
[38]	李树明, 黄治国, 罗惠波, 等. 白酒丢糟发酵生产蛋白质饲料的菌种筛选[J]. 西南大学学报:自然科学版, 2011, 33(10):27-30.
	Li SM, Huang ZG, Luo HB, et al. Microbial strain screening for liquor spent grain fermentation in protein feed production[J]. J Southwest Univ:Nat Sci Ed, 2011, 33(10):27-30.

水平 Level	因素Factors
水平 Level	A 麸皮添加量 Bran addition amount/%	B 装料量 Loading amount/g	C 初始pH Initial pH
1	5	80	3.70
2	10	100	4.67
3	15	120	5.83

水平 Level	因素Factors
水平 Level	A 麸皮添加量 Bran addition amount/%	B 装料量 Loading amount/g	C 初始pH Initial pH
1	5	80	3.70
2	10	100	4.67
3	15	120	5.83

指标 Indicator	评分标准 Scoring criteria			分数 Score
气味 Smell	无丁酸臭味,有芳香果味或明显的面包香味 With aromatic fruit or obvious bread flavor,but no butyric acid odor			14
	有微弱的丁酸臭味,或较轻的酸味,芳香味弱 With a weak butyric acid odor or a light sour odor,and weak aroma			10
	丁酸味颇重,或有刺鼻的焦糊臭或霉味 With a quite strong butyric acid odor,or a pungent burnt smell or a musty smell			4
	有较强的丁酸臭味或氨味,或几乎无酸味 With a strong butyric acid odor or an ammonia odor,or almost no acid odor			2
质地 Texture	茎叶结构保持良好 Structure of stem and leaf remains well			4
	茎叶结构保持较差 Structure of stem and leaf remains poorly			2
	茎叶结构保持极差,或有轻度的霉菌污染 Structure of stem and leaf remains extremely poor,or there is mild mold contamination			1
	茎叶腐烂或污染严重 Stems and leaves are rotted or heavily polluted			0
色泽 Tinct	与原料相似,烘干后呈淡褐色 Similar to the raw material,it is light brown after drying			2
	略有变色呈淡黄色或带褐色 Slightly discolored,light yellow or brown			1
	变色严重,墨绿色或褐色呈黄色,霉味较强 Serious discoloration,dark green or brown,yellow,and with a strong musty smell			0
总分Total score 等级Grade	16-20	10-15	5-9	0-4
总分Total score 等级Grade	优良（1级） Excellent（Level 1）	尚好（2级） Good（Level 2）	中等（3级） Medium（Level 3）	腐败（4级） Corrupt （Level 4）

指标 Indicator	评分标准 Scoring criteria			分数 Score
气味 Smell	无丁酸臭味,有芳香果味或明显的面包香味 With aromatic fruit or obvious bread flavor,but no butyric acid odor			14
	有微弱的丁酸臭味,或较轻的酸味,芳香味弱 With a weak butyric acid odor or a light sour odor,and weak aroma			10
	丁酸味颇重,或有刺鼻的焦糊臭或霉味 With a quite strong butyric acid odor,or a pungent burnt smell or a musty smell			4
	有较强的丁酸臭味或氨味,或几乎无酸味 With a strong butyric acid odor or an ammonia odor,or almost no acid odor			2
质地 Texture	茎叶结构保持良好 Structure of stem and leaf remains well			4
	茎叶结构保持较差 Structure of stem and leaf remains poorly			2
	茎叶结构保持极差,或有轻度的霉菌污染 Structure of stem and leaf remains extremely poor,or there is mild mold contamination			1
	茎叶腐烂或污染严重 Stems and leaves are rotted or heavily polluted			0
色泽 Tinct	与原料相似,烘干后呈淡褐色 Similar to the raw material,it is light brown after drying			2
	略有变色呈淡黄色或带褐色 Slightly discolored,light yellow or brown			1
	变色严重,墨绿色或褐色呈黄色,霉味较强 Serious discoloration,dark green or brown,yellow,and with a strong musty smell			0
总分Total score 等级Grade	16-20	10-15	5-9	0-4
总分Total score 等级Grade	优良（1级） Excellent（Level 1）	尚好（2级） Good（Level 2）	中等（3级） Medium（Level 3）	腐败（4级） Corrupt （Level 4）

试验号Test No.	A	B	C	试验结果Test results
试验号Test No.	A	B	C	粗蛋白含量 Crude protein content/%	粗纤维含量 Crude fiber content/%	乙偶姻含量 Acetoin content/（mg·g^-1distiller’s grains）	烟酸含量 Nicotinic acid content/（mg·g^-1distiller’s grains）
1	1	1	1	35.21	12.73	37.30	1.21
2	1	2	3	32.10	13.16	15.68	3.55
3	1	3	2	33.62	12.69	29.68	2.14
4	2	1	2	30.56	11.54	28.79	1.46
5	2	2	1	34.66	12.27	29.02	1.88
6	2	3	3	30.70	11.49	3.65	3.10
7	3	1	3	29.19	10.34	11.65	1.17
8	3	2	2	30.31	10.71	0.00	1.23
9	3	3	1	32.09	9.86	31.22	0.84
粗蛋白含量 Crude protein content/%
k1	33.64	31.65	33.99
k2	31.97	32.36	31.50
k3	30.53	32.14	30.66
R	3.11	0.71	3.33
粗纤维含量 Crude fiber content/%
k1	12.86	11.54	11.62
k2	11.77	12.05	11.64
k3	10.30	11.35	11.66
R	2.56	0.70	0.04
乙偶姻含量Acetoin content/（mg·g^-1distiller’s grains）
k1	27.55	25.91	32.51
k2	20.49	14.90	19.49
k3	14.29	21.52	10.33
R	13.26	11.01	22.18
烟酸含量Nicotinic acid content /（mg·g^-1distiller’s grains）
k1	2.30	1.28	1.31
k2	2.15	2.22	1.61
k3	1.08	2.03	2.61
R	1.22	0.94	1.30