Optimization of biological deodorization process and analysis of volatile flavor in Gracilaria lemaneiformis
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摘要:
目的 优化龙须菜微生物发酵脱腥工艺，为龙须菜的深加工开发利用提供技术支持。 方法 通过感官模糊综合评价，在筛选出生物脱腥微生物菌种组合的基础上，采用Box-Behnken响应面法优化基料白砂糖及苹果酸添加量、菌接种量、发酵时间、发酵温度等因素工艺参数。通过HS-SPME-GC/MS等技术，分析发酵前后龙须菜挥发性风味物质的变化。 结果 最优脱腥工艺：脱腥微生物菌种组合为1∶1的酿酒酵母JJ4和副干酪乳杆菌副干酪亚种RP38、接种量各5%；基料白砂糖添加量（m/m）为2.0%、苹果酸添加量（m/m）为0.2%，发酵时间68 h，发酵温度29 ℃。发酵脱腥后，具有脂肪味、腥臭味、鱼腥味的环辛醇、1-甲基环庚醇、辛醛、正壬醛、（E）-2-壬烯醛消失，具有泥土腥味中的3-辛酮含量明显降低，新增了具有芳香味的β-苯乙醇、水果香的己酸乙酯、酒香的辛酸乙酯、甜蜜香的乙酸苯乙酯、椰子香气的壬酸乙酯等。 结论 采用酿酒酵母JJ4和副干酪乳杆菌副干酪亚种RP38组合发酵龙须菜，削除了醇、醛等腥味物质，增加了β-苯乙醇及酯类等香味物质，对龙须菜有明显的脱腥赋香效果。 Abstract:Objective Optimize the microbial fermentation and deodorization process of Gracilaria lemaneiformis (GL), and provide certain technical support for the deep processing and utilization of GL. Method Through the fuzzy comprehensive evaluation of sensory, on the basis of screening out the combination of biological deodorizing microorganisms, using the Box-Behnken response surface method o logy to optimize the added amount of malic acidandsugar, inoculation amount, time of fermentation, fermentation temperature factor parameters. Analyze the changes of volatile flavor compounds in GL before and after fermentation by HS-SPME-GC/MS and other techniques. Results The optimal deodorization process is: the combination of deodorizing microbial strains is 1∶1 Saccharomyces cerevisia JJ4 and Lactobacillus paracasei subsp. Paracasei RP38;sugar added 2.0%, malic acid added 0.2%, inoculation amount 5% each, fermentation time 68 h, fermentation temperature 29℃.After fermentation and deodorization, cyclooctanol, 1-methylcycloheptanol, octanal, nonanal, 2-nonenal, (E)-which have fatty, fishy and fishy odors disappear, the content of 3-octanoneis significantly reduced, added aromatic phenylethyl Alcohol, fruity hexanoic acid, ethyl ester, wine scented octanoic acid, ethyl ester, sweet scented hexanoic acid, 2-phenylethyl ester, coconut scented nonanoic acid, ethyl ester. Conclusion The combination of JJ4 and RP38 is used to ferment GL to remove the smelly substances such as alcohol and aldehydes, and increase the flavor substances such asphenylethylalcohol and esters.Andit has obvious deodorant and fragrance effect on GL. -
图 1 不同菌种发酵对脱腥效果的影响
Figure 1. Influence of fermentation of different strains on the effect of deodorization
图 2 发酵条件对脱腥效果的影响
Figure 2. Influence of fermentation conditions on the effect of deodorization
图 3 发酵温度和发酵时间交互作用对感官评分的响应面和等高线
Figure 3. Tesponse surface and contour map of the interaction between fermentation temperature and fermentation time to sensory score
图 4 龙须菜挥发性物质相对含量对比
Figure 4. Comparison of relative content of volatile substances in GL
表 1 菌种接种方案
Table 1. Bacteria inoculation program
处理组 Treatment group CK 1 2 3 4 5 6 菌种 Strain 未接种 Blank treatment RP38 R23 JJ4 RP38+R23 RP38+JJ4 R23+JJ4 
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表 2 响应面试验设计因素与水平
Table 2. Factors and levels of response surface experimental design
因素 Factor 水平 Level −1 0 1 A：菌接种量 Inoculation amount/% 3 5 7 C：发酵温度 Fermentation temperature/℃ 27 30 33 D：发酵时间 Fermentation time/h 60 72 84
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表 3 龙须菜感官评分标准
Table 3. Sensory scoring criteria of Asparagus
项目
Items权重 Weights 优等 Excellent 良好 Good 合格 Qualified 不合格 Unqualified 腥味（0.35）
Fishy smell无腥味
Odourless腥味不明显
The smell is not obvious有腥味
Smelly腥味重
Strong fishy smell香味（0.35）
Fragrance发酵香浓郁，愉悦感强
Fermentation fragrance, a strong sense of pleasure发酵香明显，有愉悦感
Fermentation fragrance is obvious, there is a sense of pleasure发酵香较淡，一般愉悦感
Fermented fragrance is lighter, generally pleasant发酵香不明显，无愉悦感
Fermentation is not obvious, no pleasure滋味（0.25）
Taste浓厚的藻味，适口性强
Strong algal smell, palatability藻味明显，适口性较好
Obvious algae smell, good palatability藻味较淡，适口性一般
The taste of algae is light and the palatability is average藻味不明显，适口性差
Algae smell is not obvious, poor palatability色泽（0.05）
Color枣红色，无褐变
Maroon red, no browning枣红色，轻微褐变
Maroon red, slightly browned颜色较暗淡，有褐变
The color is dull, with browning颜色暗淡，褐变明显
The color is dull, browning is obvious
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表 4 各等级的轶值
Table 4. Proliferation value of each grade
项目
Items优等
Excellent良好
Good及格
Qualified不及格
Unqualified分值 Points （91～100） （71～90） （41～60） （0～40） 轶 Anecdote 90 70 50 20
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表 5 响应面试验设计与结果
Table 5. Design matrix and experiment results of responsesurface method
处理
Group菌接种量
Inoculation
amount发酵温度
Fermentation
temperature发酵时间
Fermentation
time感官评分
Sensory
score1 0 0 0 73.31 2 1 0 1 63.52 3 0 0 0 71.21 4 −1 −1 0 63.34 5 0 −1 −1 70.82 6 0 −1 1 63.4 7 0 0 0 71.11 8 0 1 1 62.44 9 −1 0. −1 61.83 10 0 0 0 71.57 11 0 0 0 72.86 12 1 −1 0 64.52 13 −1 0 1 56.81 14 1 1 0 59.24 15 1 0 −1 63.69 16 −1 1 0 59.82 17 0 1 −1 63.16
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表 6 二次回归模型方差分析
Table 6. Analysis of variance of quadratic regression model
方差来源
Source of variance平方和
Sum of square自由度
Degree of freedom均方
Mean squareF值
F valueP值
P value显著性
Significance模型 Model 423.77 9 47.09 25.18 0.0002 ** A 10.51 1 10.51 5.62 0.0495 * B 37.93 1 37.93 20.29 0.0028 ** C 22.21 1 22.21 11.88 0.0107 * AB 0.77 1 0.77 0.41 0.5404 AC 5.88 1 5.88 3.15 0.1194 BC 11.22 1 11.22 6.00 0.0441 * A2 199.72 1 199.97 106.82 ＜0.0001 ** B2 48.52 1 48.52 25.95 0.0014 ** C2 56.47 1 56.47 30.20 0.0009 ** 残差 Residual 13.09 7 1.87 失拟项 Lack of fit 9.03 3 3.01 2.97 纯误差 Pure error 4.06 4 1.01 总和 Sum 436.85 16 注：“*”表示影响显著，“**”表示影响极显著。
Note: ‘*’ Indicates significant impact,‘**’ Indicates that the impact is extremely significant.
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表 7 龙须菜挥发性风味物质HS-SPME-GC-MS分析结果
Table 7. Analysis of volatile flavor components in GL by HS-SPME-GC-MS
序号
Number保留时间
Keep
time保留指数
Retention
index化合物
Compound风味描述[20, 22]
Flavor description相对含量
Relative content %英文名称
English name中文名称
Chinese name分子式
Molecular formulaCK 处理
Group烃类
Hydrocarbon1 6.963 974 Cyclopentane, 1,1,3,3-tetramethyl- 1,1,3,3 -四甲基环戊烷 C9H18 4.42±0.20 0.66±0.16 2 7.295 984 Cyclohexane, 1-ethyl-1-methyl- 1-乙基-1-甲基-环己烷 C9H18 — 3.33±0.02 3 9.381 1045 2,6-Octadiene, 2,6-dimethyl- 2,6-二甲基-2,6-辛二烯 C10H18 花香味
Floral fragrance1.18±0.20 — 4 14.443 1186 1,3-Cyclohexadiene,1,3,5,5,6,6-hexamethyl- 1,3,5,5,6,6六甲基-1,3 -环己二烯 C12H20 果香
Fruity1.72±0.35 0.56±0.12 5 16.223 1600 Pentadecane 十五烷 C15H32 柏木等香气
Cypress and other aromas1.69±0.11 0.31±0.09 6 23.419 1734 Heptadecane 十七烷 C17H36 花果香
Floral and fruity1.07±0.18 — 7 24.903 1870 Nonadecane 正十九烷 C19H40 6.52±0.60 1.1±0.08 8 26.285 2005 Eicosane 正二十烷 C20H42 椰子香
Coconut fragrance0.41±0.21 — 9 26.377 2014 Heneicosane 正二十一烷 C21H44 0.44±0.13 — 醇类
Alcohol10 10.35 1073 Cyclooctanol 环辛醇 C8H16O 脂肪味，腥臭味
Fatty smell5.75±0.90 — 11 12.05 1121 1-Methylcycloheptanol 1-甲基环庚醇 C8H16O 青草味、脂肪味
Grassy, fatty1.16±0.18 — 12 12.297 1128 Phenylethyl Alcohol β-苯乙醇 C8H10O — 48.88±0.85 13 15.032 1204 5,7-Octadien-3-ol, 2,4,4,7-tetramethyl-, (E)- (E)-2,4,4,7-四甲基-5.7 -辛二烯-3-醇 C12H22O 臭味
Smelly— 6.38±0.05 醛类
Aldehyde14 7.683 995 Octanal 辛醛 C8H16O 鱼腥味、脂肪味
Fishy, fatty2.35±0.16 — 15 11.848 1116 Nonanal 正壬醛 C9H18O 脂肪味、柑橘味
Fat flavor, citrus flavor1.77±0.10 — 16 14.082 1177 2-Nonenal, (E)- (E)-2-壬烯醛 C9H16O 紫罗兰和黄瓜味
Violet and cucumber flavor1.83±0.17 — 17 15.563 1227 Decanal 癸醛 C10H20O 酒香
Wine flavor1.31±0.23 0.48±0.06 18 15.933 1243 1-Cyclohexene-1-carboxaldehyde, 2,6,6-trimethyl- β-环柠檬醛 C10H16O 1.74±0.09 0.64±0.13 19 17.07 1292 2-Undecenal 2-十一烯醛 C11H20O 甜蜜香
Sweet fragrance5.42±0.13 0.53±0.10 20 26.734 2045 4,8,12-Tetradecatrienal, 5,9,13-trimethyl- 5,9,13-三甲基 -4,8,12-十四碳三烯醛 C17H28O 0.45±0.11 0.67±0.02 酮类
Ketone21 7.158 980 3-Octanone 3-辛酮 C8H16O 蘑菇香、泥土腥
Mushroom smell, earthy19.73±0.17 2.39±0.17 22 10.768 1085 Furylhydroxymethyl ketone 羟甲基呋喃酮 C6H6O3 12±0.14 — 23 16.307 1259 1H-Pyrrole-2,5-dione, 3-ethyl-4-methyl- 3-乙基-4-甲基-吡咯-2,5-二酮 C7H9NO2 果香
Fruity0.59±0.12 — 24 20.904 1530 5,9-Undecadien-2-one, 6,10-dimethyl- 6,10-二甲基-5,9-十一双烯-2-酮 C13H22O 2.09±0.18 0.73±0.06 25 21.508 1576 3-Buten-2-one, 4-(2,6,6-trimethyl-1-cyclohexen-1-yl)- β-紫罗酮 C13H20O 6.61±0.07 2.68±0.01 26 26.835 2054 2-Nonadecanone 2-十九烷酮 C19H38O 1.53±0.07 — 酯类
Ester27 7.587 992 Hexanoic acid, ethyl ester 己酸乙酯 C8H16O2 果香
Fruity— 3.65±0.17 28 15.331 1217 Octanoic acid, ethyl ester 辛酸乙酯 C10H20O2 酒香
Wine flavor— 6.82±0.26 29 16.85 1283 Acetic acid, 2-phenylethyl ester 乙酸苯乙酯 C10H12O2 甜蜜香
Sweet fragrance— 1.09±0.10 30 17.845 1334 Nonanoic acid, ethyl ester 壬酸乙酯 C11H22O2 椰子香气
Coconut aroma— 0.4±0.13 31 19.758 1450 Ethyl trans-4-decenoate 反式-4-癸烯酸乙酯 C11H22O2 臭味
Bad smell— 0.98±0.45 32 19.921 1461 Decanoic acid, ethyl ester 癸酸乙酯 C12H24O2 椰子香味
Coconut fragrance— 2.79±0.29 33 20.866 1528 Octanoic acid, 3-methylbutyl ester 辛酸异戊酯 C13H26O2 椰子香味
Coconut fragrance— 0.65±0.14 34 22.381 1646 2(4H)-Benzofuranone, 5,6,7,7a-tetrahydro-4,4,7a-trimethyl-, (R)- 二氢猕猴桃内酯 C11H16O2 0.89±0.09 — 35 23.237 1718 2,2,4-Trimethyl-1,3-pentanediol diisobutyrate 2,2,4-三甲基-1,3-戊二醇双异丁酸酯 C16H30O4 0.58±0.09 1.39±0.14 36 23.297 1723 Dodecanoic acid, ethyl ester 月桂酸乙酯 C14H28O2 花果香
Floral and fruity— 0.91±0.04 37 24.071 1792 Hexanoic acid, 2-phenylethyl ester 己酸-2-苯乙酯 C14H20O2 — 0.66±0.17 38 25.668 1945 Pentadecanoic acid, ethyl ester 十五烷酸乙酯 C17H34O2 — 0.34±0.04 39 26.05 1982 Tetradecanoic acid, ethyl ester 肉豆蔻酸乙酯 C16H32O2 — 0.94±0.07 40 26.578 2032 Isopropyl palmitate 棕榈酸异丙酯 C19H38O2 0.34±0.06 — 41 27.084 2077 Dibutyl phthalate 邻苯二甲酸二丁酯 C16H22O4 2.24±0.16 0.65±0.13 42 27.488 2112 Heptadecanoic acid, ethyl ester 十七烷酸乙酯 C19H38O2 蜡香、奶油味
Wax incense, fragrant cream— 0.14±0.06 43 28.296 2176 Phthalic acid, butyl hexyl ester 邻苯二甲酸丁基己基酯 C18H24O4 0.83±0.08 0.51±0.10 44 28.431 2187 Ethyl 9-hexadecenoate 9-十六碳烯酸乙酯 C18H34O2 — 0.2±0.07 45 28.48 2191 (E)-9-Octadecenoic acid ethyl ester （E）-9-十八碳烯酸乙酯 C20H38O2 — 0.21±0.06 46 28.75 2216 Nonadecanoic acid, ethyl ester 十九烷酸乙酯 C21H42O2 — 0.47±0.18 酸类
Acid47 14.667 1192 4-Methyloctanoic acid 4-甲基辛酸 C9H18O2 2.49±0.03 — 48 17.296 1303 Nonanoic acid 壬酸 C9H18O2 2.93±0.07 0.82±0.24 49 19.399 1426 n-Decanoic acid 癸酸 C10H20O2 臭味
Bad smell0.44±0.17 5.21±0.18 50 22.804 1681 Tridecanoic acid 十三酸 C13H26O2 1.8±0.16 1.16±0.12 51 25.728 1951 Pentadecanoic acid 十五酸 C15H30O2 0.42±0.03 0.31±0.15 52 28.34 2180 Octadecanoic acid 十八酸 C18H36O2 0.93±0.05 0.35±0.02 其他
Other53 9.705 1054 Octanenitrile 正辛腈 C8H15N 花香
Potpourri1.99±0.43 — 54 22.145 1627 1,3-Benzenediol, 5-pentyl- 5-戊基间苯二酚 C11H16O2 1.92±0.12 — 注：“—”表示未检出。
Note: Means ‘—’not detected.
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