酿酒酵母高密度增殖酒糟多肽糖蜜培养基优化

苏昊; 梁璋成; 林晓姿; 何志刚; 邓冬莲; 林晓婕

doi:10.19303/j.issn.1008-0384.2021.05.016

酿酒酵母高密度增殖酒糟多肽糖蜜培养基优化

doi: 10.19303/j.issn.1008-0384.2021.05.016

苏昊^{1, 2,},
梁璋成^{1, 2},
林晓姿^{1, 2},
何志刚^{1, 2, ,},
邓冬莲^{1, 2},
林晓婕^{1, 2}

1.
福建省农业科学院农业工程技术研究所，福建　福州　350003
2.
福建省农产品（食品）加工重点实验室，福建　福州　350003

基金项目: 福建省科技计划公益类专项（2019R1032-5、2018R1014-5）

详细信息

作者简介:
苏昊（1984−），男，博士，助理研究员，主要从事食品发酵与酿造方面研究（E-mail：suhaoxp@163.com）

通讯作者:
何志刚（1964−），男，研究员，主要从事农产品贮藏与加工方面研究（E-mail：njgzx@163.com）

中图分类号: TS 261.1
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出版历程
- 收稿日期: 2020-10-15
- 修回日期: 2021-03-15
- 刊出日期: 2021-05-31

Optimized Medium Utilizing Distiller's Grains Polypeptide for High-density Culture of Saccharomyces cerevisiae

SU Hao^{1, 2
,},
LIANG Zhangcheng^{1, 2},
LIN Xiaozi^{1, 2},
HE Zhigang^{1, 2
, ,},
DENG Donglian^{1, 2},
LIN Xiaojie^{1, 2}

1.
Institute of Agricultural Engineering Technology, Fujian Academy of Agricultural Sciences, Fuzhou, Fujian　350003, China
2.
Fujian Key Laboratory of Agricultural Products (Food) Processing, Fuzhou, Fujian　350003, China

摘要

摘要: 目的研发适用于酿酒酵母菌剂生产的高密度低成本培养基。方法以酿酒酵母JH301为对象菌，以糖蜜为基础培养基，采用单因素试验筛选适宜的碳源、氮源、无机盐和天然产物增殖因子，通过U₁₆（8⁸）均匀试验优化培养基成分。结果研究筛选出适宜的碳源为棉籽糖、葡萄糖和海藻糖，氮源为胰蛋白胨，无机盐为磷酸二氢钾，天然产物增殖因子为酒糟多肽、大米糖化液和大豆多糖。获得优化培养基配方为：在可溶性固形物含量为10%的糖蜜中添加酒糟多肽23.72 g·L⁻¹、大米糖化液44.15 g·L⁻¹、大豆多糖31.88 g·L⁻¹、棉籽糖22.45 g·L⁻¹、葡萄糖13.97 g·L⁻¹、海藻糖15.00 g·L⁻¹、胰蛋白胨27.25 g·L⁻¹、磷酸二氢钾3.00 g·L⁻¹，调节pH至4.5±0.2。酵母细胞增殖培养生物量可达1.48×10⁸ cfu·mL⁻¹，比基础糖蜜培养基提高了1个数量级。结论酒糟多肽中富含多种对酿酒酵母细胞增殖起促进作用的小分子多肽成分，作为天然产物添加到糖蜜培养基中可显著提高酵母产量，是一种来源丰富、低成本的优良复合营养源，相关研究内容可为高活力酵母菌剂制备提供思路和参考依据。
- 酿酒酵母 /
- 糖蜜 /
- 酒糟多肽 /
- 高密度培养 /
- 增殖培养基
Abstract: Objective An optimized low-cost medium for high-density culture of Saccharomyces cerevisiae was formulated. Method S. cerevisiae JH301 was cultured on the basic molasses media of varied compositions to determine the optimal carbon source, nitrogen source, inorganic salt, and natural growth promoters for the yeast preparation in a single factor test. Optimization models using different statistical analyses were compared for the formulation finalization. Result From a U₁₆(8⁸) uniform experiment, the optimized medium formulation was determined to apply the carbon source that included glucose, cottonseed sugar, and trehalose, tryptone for the nitrogen source, K₂HPO₄ as the inorganic salt, and the natural growth promoters that consisted of distiller's grains polypeptide, rice saccharification solution, and soybean polysaccharide. The finalized formula called for 23.72 g·L⁻¹ distiller's grains polypeptide, 44.15 g·L⁻¹ rice saccharification solution, 31.88 g·L⁻¹ soybean polysaccharide, 22.45 g·L⁻¹ cottonseed sugar, 13.97 g·L⁻¹ glucose, 15.00 g·L⁻¹ trehalose, 27.25 g·L⁻¹ tryptone, and 3.00 g·L⁻¹ K₂HPO_4, in addition to 10% soluble solids molasses. The resulting yeast biomass reached 1.48×10⁸ cfu·mL⁻¹, which was one order of magnitude higher than the culture obtained from the basic molasses medium. Conclusion Rich in small molecular polypeptides capable of promoting the cell proliferation of S. cerevisiae, the natural distiller's grains polypeptide was formulated in the medium to yield a low-cost preparation for the yeast.
- Saccharomyces cerevisiae /
- molasses /
- distiller's grains polypeptide /
- high-density culture /
- proliferation medium

HTML全文

图 1 碳源种类及质量浓度对JH301生长的影响

注：图中不同小写字母表示同一处理不同质量浓度之间差异显著（P＜0.05），不同大写字母表示不同处理组中最高OD₆₀₀增量组间的差异显著（P＜0.05）。图2～4同。

Figure 1. Effects of C sources and concentrations on growth of JH301

Note: Data with different lowercase letters indicate significant differences between different concentrations under same treatment; data with different capital letters, significant differences on highest OD₆₀₀ increment group between different treatments (P＜0.05). Same for Fig.2–4.

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图 2 氮源种类及质量浓度对JH301生长的影响

Figure 2. Effects of N sources and concentrations on growth of JH301

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图 3 无机盐种类及质量浓度对JH301生长的影响

Figure 3. Effects of inorganic salts and concentrations on growth of JH301

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图 4 天然产物增殖因子及质量浓度对JH301生长的影响

Figure 4. Effects of natural growth promoters and concentrations on growth of JH301

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表 1 U₁₆（8⁸）均匀试验结果

Table 1. Results of U₁₆(8⁸) uniform test

因子 Factor	x₁磷酸二氢钾 KH₂PO₄ / （g·L⁻¹）	x₂胰蛋白胨 Tryptone/ （g·L⁻¹）	x₃海藻糖 Trehalose/ （g·L⁻¹）	x₄葡萄糖 Glucose/ （g·L⁻¹）	x₅棉籽糖 Raffinose/ （g·L⁻¹）	x₆酒糟多肽（Distiller's grains polypeptide, DGP）/ （g·L⁻¹）	x₇大豆多糖（Soybean polysaccharide, SP）/ （g·L⁻¹）	x₈大米糖化液（Rice saccharification solution, RSS）/ （g·L⁻¹）	菌体密度对数值 Cell density
N1	5.5	10	55	10	30	40	20	60	7.29
N2	4.5	45	55	10	5	5	20	40	7.19
N3	5.5	30	45	30	15	10	40	65	7.44
N4	6.0	15	35	25	10	25	10	40	7.50
N5	4.5	20	75	30	40	35	40	45	7.58
N6	4.0	40	35	5	35	25	30	65	7.51
N7	3.0	15	65	5	10	20	45	50	7.71
N8	5.0	25	15	0	40	15	15	45	7.57
N9	4.0	25	15	35	5	30	25	55	7.60
N10	3.5	10	25	20	35	10	35	35	7.64
N11	5.0	35	85	0	15	30	35	35	7.86
N12	3.0	30	45	15	20	40	15	30	7.57
N13	6.0	40	65	35	35	20	25	30	7.48
N14	6.5	20	85	15	20	5	30	55	7.62
N15	6.5	45	25	20	25	35	45	50	7.59
N16	3.5	35	75	25	25	15	10	60	7.47

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表 2 模型1优选培养基组合

Table 2. Medium formulation optimization by stepwise regression of multiple factors and square terms

成分 Factors	磷酸二氢钾 KH₂PO₄	胰蛋白胨 Tryptone	海藻糖 Trehalose	葡萄糖 Glucose	棉籽糖 Raffinose	酒糟多肽 DGP	大豆多糖 SP	大米糖化液 RSS
水平编码值 Level	1.00	4.45	1.00	3.79	4.49	4.74	5.38	3.83
质量浓度 Concentration/（g·L⁻¹）	3.00	27.25	15.00	13.97	22.45	23.72	31.88	44.15

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表 3 模型2优选培养基组合

Table 3. Medium formulation optimization by stepwise regression of multiple factors and interaction

成分 Factors	磷酸二氢钾 KH₂PO₄	胰蛋白胨 Tryptone	海藻糖 Trehalose	葡萄糖 Glucose	棉籽糖 Raffinose	酒糟多肽 DGP	大豆多糖 SP	大米糖化液 RSS
水平编码值 Level	8.00	8.00	1.00	8.00	8.00	8.00	1.00	8.00
质量浓度 Concentration/（g·L⁻¹）	6.50	45.00	15.00	35.00	40.00	40.00	10.00	65.00

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表 4 模型3优选培养基组合

Table 4. Medium formulation optimization by quadratic polynomial stepwise regression analysis

成分 Factors	磷酸二氢钾 KH₂PO₄	胰蛋白胨 Tryptone	海藻糖 Trehalose	葡萄糖 Glucose	棉籽糖 Raffinose	酒糟多肽 DGP	大豆多糖 SP	大米糖化液 RSS
水平编码值 Level	8.00	8.00	1.00	8.00	8.00	8.00	1.00	8.00
质量浓度 Concentration/（g·L⁻¹）	6.50	45.00	15.00	35.00	40.00	40.00	10.00	65.00

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