酶解联合高温处理对可可粉风味的影响

钱蕾; 于海燕; 娄新曼; 田怀香

doi:10.13386/j.issn1002-0306.2021120085

酶解联合高温处理对可可粉风味的影响

doi: 10.13386/j.issn1002-0306.2021120085

钱蕾^{1, 2,},
于海燕¹,
娄新曼¹,
田怀香^1, ,

1.
上海应用技术大学香料香精技术与工程学院，上海 201418
2.
上海牡丹香精香料有限公司，上海 201210

详细信息

作者简介:
钱蕾（1991−），女，硕士研究生，研究方向：天然提取和香原料开发，E-mail：lilyqian0919@126.com

通讯作者:
田怀香（1976−），女，博士，教授，研究方向:食品风味化学，E-mail：tianhx@sit.edu.cn

中图分类号: TS274
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- 文章访问数: 11
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出版历程
- 收稿日期: 2021-12-08
- 网络出版日期: 2022-08-05

Effect of Enzymatic Hydrolysis Combined with High Temperature Processing on the Flavor of Cocoa Powder

QIAN Lei^{1, 2
,},
YU Haiyan¹,
LOU Xinman¹,
TIAN Huaixiang^{1
, ,}

1.
School of Perfume and Aroma Technology, Shanghai Institute of Technology, Shanghai 201418, China
2.
Shanghai Peony Flavor & Fragrance Co., Ltd., Shanghai 201210, China

摘要

摘要: 本文考察了酶解和高温联合处理对可可风味的作用。首先以游离氨基酸含量为评价指标，研究了5种蛋白酶对脱脂可可粉酶解液中游离氨基酸的影响，并进一步采用顶空-气相色谱-质谱联用（headspace gas chromatography mass spectrometry，HS-GC-MS）结合香气活力值（odor activity value，OAV）以及感官评价研究了高温（90~180 ℃）对可可挥发性物质的作用。结果表明，酶解处理提高了总氨基酸含量，为酶解前的1.4~5倍，且与未经酶处理的对照样（12种挥发物）相比，5种联合处理的样品分别检测出29（天冬氨酸酶）、31（酸性蛋白酶）、29（风味蛋白酶）、30（菠萝蛋白酶）、32（木瓜蛋白酶）种挥发物，挥发性物质种类明显增加。当反应温度达到150 ℃以上更有助于可可中具有坚果和奶油香气的杂环化合物的生成；由感官评价可得，经风味蛋白酶处理并在150 ℃反应2 h的可可风味最佳。基于OAV与感官评价的偏最小二乘（partial least squares，PLS）模型验证可得：酶解和高温处理使可可在保持原有巧克力香气的同时增加花果香和奶香风味，有助于提升可可整体香气品质。
- 可可粉 /
- 酶解 /
- 高温处理 /
- 美拉德反应 /
- 氨基酸 /
- 可可风味 /
- 顶空-气相色谱-质谱联用(HS-GC-MS) /
- 感官评价
Abstract: The study aimed to evaluate the effect of enzymatic hydrolysis and high temperature on the flavor of cocoa. First, the effect of five proteases on free amino acids in enzymatic hydrolysate were investigated. Subsequently, headspace gas chromatography mass spectrometry (HS-GC-MS), odor activity value (OAV) and sensory valuation were applied to explore the influence of high temperature (90~180 ℃) on the volatile compounds of cocoa. The results showed that amino acids content increased by 1.4~5.0 times after enzymatic hydrolysis, and compared with the control sample untreated by enzyme (12 volatiles), five combinatorial treated samples promoted volatiles, which were identified 29 (asparaginase)、31 (acid Protease)、29 (flavourzyme)、30 (bromelain)、32 (papain) volatiles, respectively. More heterocyclic compounds with nutty and creamy aroma were generated when the temperature was up to 150 ℃, and the sensory valuation indicated that flavor protease hydrolysis combined with 150 ℃ heating for 2 h presented the best flavor characteristics. Meanwhile, partial least squares (PLS) analysis model based on OAV and sensory evaluation showed that enzymatic hydrolysis combined with high temperature gave cocoa more flower, fruit and milk aroma while still maintained chocolate flavor, confirming its contribution to flavor promotion.
- cocoa powder /
- enzymatic hydrolysis /
- high temperature processing /
- Maillard reaction /
- amino acid /
- cocoa flavor /
- headspace-gas chromatography mass spectrometry (HS-GC-MS) /
- sensory valuation

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图 1 5种蛋白酶与可可粉酶解后氨基酸含量变化关系

Figure 1. Changes in the concentration of free amino acid from five enzymic hydrolysates

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图 2 不同反应温度的可可粉酶解液挥发物变化图

Figure 2. Changes of cocoa enzymic hydrolysates volatile compounds at different temperature

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图 3 不同反应温度对可可挥发物OAV的聚类树图

Figure 3. Tree graph of different temperature on the volatile OAV of cocoa

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图 4 可可联合处理反应物的感官评价

Figure 4. Sensory evaluation on enzymatic hydrolysis combined with high temperature of cocoa

注：*表示存在显著性差异（P<0.05）；**表示存在极显著性差异（P<0.01）。

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图 5 不同酶处理样品与感官属性的PLS图

Figure 5. Partial least squares regression of cocoa samples and sensory indicators

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表 1 不同酶的酶解条件

Table 1. Enzymatic hydrolysis conditions of different enzymes

蛋白酶	简写	酶活度（kU·g⁻¹）	酶解pH	酶解温度（℃）	酶添加量（kU·g⁻¹）
天冬氨酸酶	Asp	100	2.0	40	10
酸性蛋白酶	Acid	500	3.5	40	10
风味蛋白酶	Fla	300	6.4	55	10
菠萝蛋白酶	Bro	1000	6.4	55	10
木瓜蛋白酶	Pap	1000	6.4	55	10

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表 2 感官描述词和评分

Table 2. Sensory evaluation descriptions

	描述词	感官描述	参比物
嗅觉	巧克力（10分）	巧克力的特殊香气	α-异丁基苯乙醇溶液20 mg/L
	奶香（10分）	新鲜牛乳的特征风味	0.1 mg/kg δ-十二烷内酯
	焦糖（10分）	一种带有甜香、焦苦的气味	糠醇水溶液60 mg/L
	花香（10分）	鲜花特有的复合香气、带有香甜感	香叶醇水溶液1 mg/L
	果香（10分）	熟透水果浓郁香气，甜香花香合感	苹果
	坚果（10分）	烤味、木质味、油味的混合	2,6-二甲基吡嗪2 mg/L醇水溶液
	木香（10分）	新切割的木材，树皮味	10 mg/L α-蛇麻烯
	辛香（10分）	五香八角的气味	五香八角香料
	烟熏（10分）	烟气熏制食品过程产生的一类特殊风味	10 g/L香芹酚丙二醇溶液
味觉	甜（10分）	如糖似蜜	蔗糖质量浓度为5.0 g/500 mL）
	酸（10分）	像醋的味道	柠檬酸质量浓度为0.25 g/500 mL）
	涩（10分）	舌头口腔表面收缩、收敛感	1 g/L的单宁酸水溶液
	苦（10分）	苦瓜、啤酒、咖啡等特有的味感	氯化奎宁质量浓度为0.072 g/500 mL）
	油脂（10分）	植物油、矿物油的气味	植物调和油

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表 3 可可联合处理反应物的挥发物HS-GC-MS分析

Table 3. Contents of volatile compounds from Cocoa combined reactants by HS-GC-MS

化合物	RI		鉴别方式	阈值	香气描述	对照样	天冬氨酸酶	酸性蛋白酶	风味蛋白酶	菠萝蛋白酶	木瓜蛋白酶
化合物	文献值	计算值	鉴别方式	μg/g	香气描述	挥发物质量分数（μg/g）
甲酸	543	538	MS, RI	0.046	强烈的刺激性	−	10.8±0.78^b	12.56±2.38^b	77.77±73.30^a	−	7.98±2.28^b
2,3-丁二酮	600	615	MS, RI	0.005	稀释奶油	6.67±0.40^b	13.07±0.97^ab	7.64±1.80^b	22.43±3.66^a	1.77±0.41^b	3.15±0.91^b
乙酸乙酯	610	626	MS, RI	5	菠萝果香、白兰地	−	−	2.77±0.55^b	158.91±7.75^a	3.43±0.70^b	−
异丁醇	622	634	MS, RI	3.3	透发的酒样香气	7.62±0.51^b	−	−	29.84±1.46^a	6.39±1.36^b	−
乙酸	646	644	MS, RI	10000	强烈的酸	305.41±21.42^b	2128.74±697.42^a	1948.8±77.17^a	1133.43±239.48^a	14.62±3.02^b	17.20±12.74^b
丙二酸	−	675	MS, RI	−	−	−	−	−	−	0.22±0.08	0.26±0.13
异戊醛	655	650	MS, RI	−	辛辣、苹果杏仁	1.53±0.03^b	−	3.32±0.59^b	23.54±23.75^a	5.42±1.32^b	6.73±1.79^b
戊醛	658	660	MS, RI	0.012-0.1	发酵面包、坚果	−	−	1.36±0.24^b	−	0.49±0.11^b	14.71±4.32^a
2-甲基丁醛	661	661	MS, RI	10	咖啡和可可	−	−	−	32.67±12.71^a	2.12±0.50^b	2.44±1.20^b
2-戊酮	686	687	MS, RI	0.07	酒和丙酮样透发	−	−	−	−	0.36±0.10	−
羟基丙酮	688	681	MS, RI	−	−	25.17±1.69^b	154.23±3.98^a	129.46±42.92^a	53.53±64.19^b	−	12.52±3.65^b
戊二酮	696	686	MS, RI	0.07-0.1	−	6.42±0.38^b	−	−	13.47±0.66^a	−	−
3-羟基-2-丁酮	720	705	MS, RI	10000	奶油	−	−	−	20.88±0.96^a	2.22±0.46^b	20.23±5.89^a
丁酸甲酯	724	714	MS, RI	0.001-0.043	苹果和干酪	−	−	−	−	−	0.22±0.06
3-甲基丁醇	726	733	MS, RI	0.25-4.1	威士忌特征的辛辣	−	−	−	−	5.07±1.08^a	1.10±0.32^b
2-甲基丁醇	733	739	MS, RI	0.0017	水果和酒精	−	−	−	−	2.03±0.43^a	0.28±0.08^b
二甲基二硫醚	761	764	MS, RI	0.0012	洋葱	3.37±0.45^b	3.84±0.58^b	4.87±0.93^b	13.11±0.64^a	−	−
己醛	799	795	MS, RI	0.0228	青草、苹果	−	−	−	−	0.25±0.06	0.21±0.05
羟基丁酮	803	796	MS, RI	−	咖啡、奶油、香甜	−	17.99±0.38	−	−	−	−
2,3-丁二醇	803	808	MS, RI	−	甜巧克力	−	38.92±6.59^b	10.23±1.94^b	491.92±65.69^a	23.10±9.17^b	24.83±5.52^b
糠醛	818	828	MS, RI	0.28-8	面包发酵、焦糖	93.73±6.59^b	268.99±22.86^ab	250.12±61.51^ab	431.13±40.50^a	26.00±8.81^b	19.22±11.14^b
异戊酸	834	840	MS, RI	2.8	腐臭的、奶酪	−	−	−	8.14±5.16	−	−
糠醇	865	863	MS, RI	1.0-2.0	发酵、奶油、焦糖	−	12.75±0.11^b	22.61±4.31^a	−	−	−
环戊烯二酮	880	880	MS, RI	−	−	−	2.75±0.47^a	1.85±0.35^ab	−	−	0.22±0.06^b
2-庚酮	889	884	MS, RI	160	水果、辛香	−	−	−	−	−	0.27±0.08
2-乙酰基呋喃	910	902	MS, RI	10	甜杏仁坚果、烟熏	−	47.35±2.11^ab	42.21±14.13^ab	133.79±6.55^a	6.22±1.31^b	3.21±0.94^b
2,6-二甲基吡嗪	916	908	MS, RI	1.5	坚果、可可、咖啡	−	0.55±0.02^b	0.37±0.06^b	0.98 ±0.28^a	0.47 ±0.11^b	0.45 ±0.08^b
己酸甲酯	936	926	MS, RI	0.087	菠萝	−	2.86±0.05^b	1.34±0.27^b	16.82±10.64^a	0.32±0.09^b	0.28±0.07^b
甲基糠醇	953	940	MS, RI	−	焦糖	−	3.21±0.07^a	1.63±0.31^b	−	−	−
5-甲基糠醛	957	954	MS, RI	6	焦糖的香甜	−	2.47±0.05^b	5.18±1.06^a	−	−	−
苯甲醛	964	969	MS, RI	0.33	苦杏仁	3.11±0.16^b	3.99±3.38^b	2.77±0.51^b	15.45±0.74^a	1.92±0.46^b	1.46±0.40^b
乙酸糠醇酯	998	991	MS, RI	50	发酵奶油、焦糖	−	2.95±0.55^a	1.09±0.22^b	−	−	−
2-乙基-6-甲基吡嗪	997	993	MS, RI	0.1	可可、坚果、烤土豆	−	0.18 ±0.19^b	0.19 ±0.19^b	0.46 ±0.15^a	0.47 ±0.04^a	0.28 ±0.28^ab
2-乙基-3-甲基吡嗪	1003	997	MS, RI	2	玉米花、坚果	−	0.24 ±0.04	0.24 ±0.02	−	0.16 ±0.07	−
苯乙醛	1045	1038	MS, RI	0.004	强烈风信子花香	2.87±0.03^b	4.42±2.92^b	12.28±2.49^ab	31.70±2.42^a	1.77±0.77^b	2.31±0.63^b
4-羟基-2,5-二甲基- 3（2H）-呋喃酮	1072	1069	MS, RI	0.06	烤面包、焦糖	−	2.35±0.07^a	2.58±0.52^a	−	−	0.40±0.10^b
庚酸	1071	1081	MS, RI	10.4	酸臭、汗臭、脂肪味	2.67±0.04^c	124.22±34.65^a	44.77±10.73^b	41.70±2.04^b	7.85±1.68^c	7.16±2.10^c
2,3,5,6－四甲基吡嗪	1087	1080	MS, RI	1	坚果、可可、奶油	−	1.80±1.08^a	0.47 ±0.17^b	0.14 ±0.10^b	−	0.21 ±0.20^b
庚酸乙酯	1108	1096	MS, RI	−	菠萝	−	−	−	−	0.38±0.07	−
苯乙醇	1121	1108	MS, RI	3.5	玫瑰花香	14.42±0.98^a	2.83±0.54^c	1.36±0.26^c	6.94±0.33^b	9.53±4.02^b	2.44±0.71^c
乙基环戊烯醇酮	1140	1145	MS, RI	500	焦糖、烟熏、咖啡	−	2.70±0.05	−	−	−	−
2,3-二氢-3,5二羟基- 6-甲基- 4（H）-吡喃-4-酮	1154	1161	MS, RI	−	焦糖样甜香	−	−	1.28±0.23^b	8.33±0.92^a	−	−
辛酸	1186	1178	MS, RI	1-19	果香	−	18.63±1.53^bc	40.02±8.06^b	80.41±34.93^a	2.74±0.70^c	1.35±0.36^c
5-羟甲基糠醛	1233	1224	MS, RI	−	黄油、焦糖	−	8.97±2.69^a	1.64±0.33^b	−	−	−
癸酸	1380	1374	MS, RI	2.2-102	蜡、水果	−	4.71±0.44^b	5.55±1.12^b	14.63±9.26^a	1.38±0.36^bc	0.40±0.10^c
月桂酸	1562	1566	MS, RI	100000	乳脂香	−	8.22±6.32^b	13.93±2.71^b	388.66±246.19^a	23.99±0.18^b	9.44±2.42^b
肉豆蔻酸	1761	1749	MS, RI	10	−	−	−	−	15.16±9.60^a	0.26±0.06^b	0.58±0.16^b
咖啡因	1840	1857	MS, RI	29	−	−	−	−	62.03±39.31^a	1.04±0.27^b	0.79±0.22^b
注： - 未查到文献值或未检出。相关数据根据webbook.nist.gov和https://www.chemicalbook.com及文献报道^{[42- 43]}查得。同行不同字母表示显著性差异，P < 0.05。

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