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抗食品腐败真菌的乳酸菌筛选、复配及混合发酵工艺优化

杜莹瑞 李市场 冯腾柱 张亚娟 陈妍 龚明贵

杜莹瑞,李市场,冯腾柱,等. 抗食品腐败真菌的乳酸菌筛选、复配及混合发酵工艺优化[J]. 食品工业科技,2022,43(12):124−131. doi:  10.13386/j.issn1002-0306.2021090312
引用本文: 杜莹瑞,李市场,冯腾柱,等. 抗食品腐败真菌的乳酸菌筛选、复配及混合发酵工艺优化[J]. 食品工业科技,2022,43(12):124−131. doi:  10.13386/j.issn1002-0306.2021090312
DU Yingrui, LI Shichang, FENG Tengzhu, et al. Screening, Compounding and Mixed Fermentation Process Optimization of Lactic Acid Bacteria against Food Spoilage Fungi[J]. Science and Technology of Food Industry, 2022, 43(12): 124−131. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021090312
Citation: DU Yingrui, LI Shichang, FENG Tengzhu, et al. Screening, Compounding and Mixed Fermentation Process Optimization of Lactic Acid Bacteria against Food Spoilage Fungi[J]. Science and Technology of Food Industry, 2022, 43(12): 124−131. (in Chinese with English abstract). doi:  10.13386/j.issn1002-0306.2021090312

抗食品腐败真菌的乳酸菌筛选、复配及混合发酵工艺优化

doi: 10.13386/j.issn1002-0306.2021090312
基金项目: 国家自然科学基金面上项目(31870093)。
详细信息
    作者简介:

    杜莹瑞(1998−),女,硕士研究生,研究方向:生物学,E-mail:duyingrui980@163.com

    通讯作者:

    李市场(1978−),男,博士,副教授,研究方向:发酵工程、酶工程和微生物育种,E-mail:lscgam@sina.com

  • 中图分类号: Q93-3

Screening, Compounding and Mixed Fermentation Process Optimization of Lactic Acid Bacteria against Food Spoilage Fungi

  • 摘要: 应用化学防腐剂控制食品中腐败微生物,防腐剂残留对人体具有一定的潜在性危害。本文筛选抗食品腐败真菌的乳酸菌,并对其进行复配及混合发酵以提高其发酵液抗真菌活性。通过24孔板双层琼脂法及分级抑菌浓度法从多株乳酸菌和丙酸杆菌中筛选抗真菌较强的菌株并确定混合菌最优组合。通过Plackett-Burman试验,最陡爬坡试验,对其发酵工艺进行优化,用96孔板酶标仪法测其发酵上清液的抗真菌活性。结果表明:最优混合菌组合为植物乳杆菌L9和费氏丙酸杆菌D5;发酵培养基优化配方:葡萄糖55 g/L,碳酸钙6.7 g/L,酵母浸粉14.8 g/L,磷酸氢二钾 0.25 g/L,硫酸锰 0.1 g/L,乙酸钠 5.0 g/L,柠檬酸铵 2.0 g/L,接种比例为5:1(D5:L9),发酵温度为37 ℃。对筛选出的L9和D5在优化后的发酵培养基进行混合发酵验证,其发酵液抗真菌活性可高达47.07 AU。
  • 图  1  6株菌株对桔青霉Q05(A)和黄曲霉Y03(B)的抑制效果

    Figure  1.  Inhibitory effects of 6 strains on Penicillium citrinum Q05 (A) and Aspergillus flavus Y03 (B)

    注:A:桔青霉Q05;B:黄曲霉Y03;乳酸乳球菌L8、L9、沃纳利葡萄球菌L7、植物乳杆菌3103、3014、3035

    图  2  乳酸菌和丙酸杆菌组合对黄曲霉Y03的抑制效果

    Figure  2.  Inhibitory effect of Lactobacillus and Propionibacterium combination on Aspergillus flavus Y03

    注:每横行6个孔中为两种不同菌株组合的三个平行。

    图  3  FIC法测定抗真菌活性

    Figure  3.  Antifungal activity assayed by FIC method

    图  4  包含12个因素标准化效应值的帕累托图

    Figure  4.  Pareto chart of standardized effects of 12 factors

    图  5  最陡爬坡试验结果

    Figure  5.  Results of the steepest climbing test

    表  1  Plackett-Burmen实验设计水平范围

    Table  1.   Range of different factors investigated of Plackett-Burman design

    编码 因素水平
    1−1
    X1 葡萄糖8040
    X2 碳酸钙3216
    X3 酵母浸粉100
    X4 酪蛋白胨100
    X5 磷酸氢二钾0.50
    X6 硫酸镁0.20
    X7 硫酸锰0.20
    X8 氯化钠20.5
    X9 接种比(D5:L9)1:55:1
    X10 培养温度3833
    X11 乙酸钠155
    X12 柠檬酸铵62
    下载: 导出CSV

    表  2  最陡爬坡试验设计

    Table  2.   Design of the steepest climbing test

    编号葡萄糖(g/L)碳酸钙(g/L)酵母浸粉(g/L)
    1401610
    24512.911.6
    3509.813.2
    4556.714.8
    5603.616.4
    6650.518
    下载: 导出CSV

    表  3  各抗真菌菌株对5种霉菌的抑制效果

    Table  3.   Inhibitory activity of antifungal strains against 5 kinds of molds

    菌株编号黑曲霉H01黄曲霉Y03枝孢霉H04桔青霉Q05交链孢霉09
    L1+++++++++++
    L2++++++
    L3+++++++++
    L4+++++++++
    L5++++++
    L6+++++++++
    L8+++
    L9+++++++++++++
    3013++++++++++++
    3014+++++++++
    3035+++++++++++++
    D1+++++++++++++
    D2+++++++++++++++
    D4++++++++++++++
    D5+++++++++++++++
    注:“+++”表示霉菌生长被完全或接近完全抑制;“++”表示霉菌生长被明显抑制,但仍有少量气生菌丝的生长;“+”表示霉菌生长受到弱的抑制;“−”表示无抑制作用。
    下载: 导出CSV

    表  4  乳酸菌和丙酸杆菌的组合对黄曲霉Y03的抑制效果

    Table  4.   Inhibitory effect of Lactobacillus and Propionibacterium combination on Aspergillus flavus Y03

    菌株D1D2D4D5
    +++++++++++
    Lactobacillus rhamnosus L1+++++++++
    Lactobacillus paracacei L2++++++++
    Lactobacillus rhamnosus L3++++++++
    Lactobacillus rhamnosus L4++++++++++
    Lactobacillus paracacei L5++++++
    Lactobacillus paracacei L6++++++
    Lactobacillus paracacei L8+++++
    Lactobacillus plantarum L9++++++++++++
    Lactobacillus plantarum 3013+++++++++
    Lactobacillus plantarum 3014+++++++++
    Lactobacillus plantarum 3035+++++++++++++
    注:表中“+++”表示霉菌生长被完全或接近完全抑制;“++”表示霉菌生长被明显抑制,但仍有少量气生菌丝的生长;“+”表示霉菌生长受到弱的抑制;“−”表示无抑制作用。
    下载: 导出CSV

    表  5  Plackett-Burman试验设计和结果

    Table  5.   Plackett-Burman experimental design and results

    实验号 ABCDEFGHJKLM抗真菌
    活性(AU)
    1−11−11111−1−111−11.20
    2111−1−111−111−1−113.11
    3−11−11−11111−1−111.60
    4−1−11−11−11111−1−135.77
    5−1−1−11−11−11111−12.10
    61−1−1−1−11−11−11113.92
    7−1−1−1−11−11−111112.60
    81−1−111−111−1−1−1−115.92
    9−11111−1−111−1110.00
    101−11111−1−111−1129.26
    111−111−1−1−1−11−11−118.49
    12−1−1−1−1−1−1−1−1−1−1−1−12.00
    13−111−111−1−1−1−11−11.74
    1400000000000018.40
    1500000000000018.09
    161−11−11111−1−11124.50
    17−1−111−111−1−1−1−110.07
    18−111−1−1−1−11−11−111.56
    1911−1−1−1−11−11−1112.17
    2011−1+1−1−1−1−11−1110.98
    2111−1−111−111−1−1−12.30
    221111−1−111−111−117.04
    2300000000000016.13
    下载: 导出CSV

    表  6  Plackett-Burman试验的方差分析

    Table  6.   Analysis of variance in Plackett-Burman design

    来源自由度Adj SSAdj MSFP
    模型132019.12155.3173.600.030*
    线性121842.59153.5493.560.032*
    葡萄糖1396.44396.4419.180.014*
    碳酸钙1343.90343.9017.970.020*
    酵母浸粉1468.20468.20110.840.009**
    酪蛋白胨12.442.4400.060.817
    磷酸氢二钾1193.58193.5784.480.063
    硫酸镁135.7735.7650.830.386
    硫酸锰186.6386.6262.010.190
    氯化钠126.6126.6110.620.453
    接种比140.4940.4930.940.358
    发酵温度1118.83118.8352.750.131
    乙酸钠175.2675.2651.740.219
    柠檬酸铵154.4354.4331.260.291
    弯曲1176.54176.5374.090.074
    误差9388.5843.176
    失拟7385.5555.07936.390.270
    纯误差23.031.514
    合计222407.70
    注:*表示显著(P<0.05),**表示极显著(P<0.01)。
    下载: 导出CSV
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出版历程
  • 收稿日期:  2021-09-28
  • 网络出版日期:  2022-04-28
  • 刊出日期:  2022-06-08

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    《食品工业科技》青年编委专栏征稿 | 杂粮与主粮复配的营养学基础