CN113558199A - 一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法 - Google Patents
一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法 Download PDFInfo
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- CN113558199A CN113558199A CN202110787705.2A CN202110787705A CN113558199A CN 113558199 A CN113558199 A CN 113558199A CN 202110787705 A CN202110787705 A CN 202110787705A CN 113558199 A CN113558199 A CN 113558199A
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Abstract
一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,将蓝蛤肉用风味蛋白酶和复合蛋白酶,在250Mpa条件下高压处理1h后,进行酶解,制得蓝蛤肉酶解冻干粉;以蓝蛤肉酶解冻干粉、D‑木糖和L‑半胱氨酸为原料,搅拌均匀后,加入去离子水,调pH值至7.0,置于油浴具塞容器中,密封,油浴反应,得到蓝蛤肉酶解物风味抗氧化剂;壳聚糖和阿拉伯胶混合,加入蓝蛤肉酶解物风味抗氧化剂,在室温下搅拌、均质,然后喷雾干燥,得到调味基料。以蓝蛤为原材料,选材天然、营养、健康,制备的调味基料无苦腥味等不良口味,具有较好的热稳定性和抗氧化性,可以作为火锅底料调味基料使用,使火锅鲜味持久,且延长火锅底料储存期。
Description
技术领域
本发明涉及一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,特别涉及一种持香型具有抗氧化功能的耐高温火锅底料蓝蛤调味基料的制备方法。
背景技术
调味品是指在饮食、烹饪和食品加工中广泛应用的,用以调和滋味和气味并具有去腥、除膻、增香、增鲜等作用的产品。调味简单地说就是调和滋味,是把各种调味料依照不同的性能和作用进行配比,通过加工工艺复合到一起达到所要求的风味。调味料的配制原理就是根据各种呈味物质和香味物质的特点、各种味觉和嗅觉现象,取得风味和口味的平衡。
火锅是中华餐饮的一种形式,其历史悠久、源远流长,并作为中华饮食的文化之一,传遍大江南北,走向世界各地。火锅调味料也得益于火锅的大发展,而成为调味料行业中的一个重要类别;同火锅相似,煲汤、麻辣烫、关东煮等产品也都需要长时间的加热蒸煮,但是传统的鸡精味精等调味料,由于长时间的高温蒸煮可能会产生焦谷氨酸钠,对健康不利;同时,传统的调味料在一定的时间内鲜味就会完全释放,随着时间的延长汤底也就失去了原有的鲜味。这些都对调味料提出了更高的要求,既能提供足够的鲜味等滋味成分,又可以长时间的不间断的释放鲜味等,还可以耐受高温,高温对其无破坏作用甚至不会产生有害物质。而且,火锅调味料中一般都含有大量的油脂性物质,油脂储存过程中容易氧化,带有哈喇味,影响火锅调味料的风味。
随着社会的发展,消费者对调味料的需要也不仅仅局限在调味上面,还希望调味品能够具有更好的“营养性”和“功能性”。蓝蛤(Aloididae aloidi)是一种小型低值贝类,富含优质蛋白和鲜味物质,因此是制备海鲜调味料的理想原料,但较重的苦腥味严重限制了蓝蛤的进一步开发应用。
发明内容
本发明要解决的技术问题是提供一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,以蓝蛤为原材料,选材天然、营养、健康,制备的调味基料无苦腥味等不良口味,具有较好的热稳定性和抗氧化性,可以作为火锅底料调味基料使用,使火锅鲜味持久,且延长火锅底料储存期。
本发明的技术方案是:
一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其具体步骤如下:
(1)超高压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤肉用风味蛋白酶和复合蛋白酶酶解,在250Mpa条件下高压处理1h后,进行酶解,制得蓝蛤肉酶解冻干粉;
(2)酶解冻干粉-木糖-半胱氨酸混合液的配制
以蓝蛤肉酶解冻干粉、D-木糖和L-半胱氨酸为原料,按照质量比5:2:1.5混合,搅拌均匀后,加入去离子水,调pH值至7.0,得到酶解冻干粉-木糖-半胱氨酸混合液;
(3)蓝蛤肉酶解物风味抗氧化剂的制备
将酶解冻干粉-木糖-半胱氨酸混合液置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂;
(4)蓝蛤肉酶解物风味抗氧化剂热稳定性处理
a.壳聚糖-阿拉伯胶-风味抗氧化剂乳液的制备
壳聚糖和阿拉伯胶按照质量比9:1混合,加入蓝蛤肉酶解物风味抗氧化剂,其中,壳聚糖和阿拉伯胶总加入质量占的蓝蛤肉酶解物风味抗氧化剂质量的5%,在室温下以2000r/min搅拌1h,制得壳聚糖-阿拉伯胶-风味抗氧化剂乳液;
b.喷雾干燥
将壳聚糖-阿拉伯胶-风味抗氧化剂乳液在9000r/mi n条件下均质10min,然后喷雾干燥,得到目标产物具有抗氧化功能的耐高温蓝蛤调味基料。
进一步的,喷雾干燥条件为:进料流量9mL/min,进风温度为145℃,出风温度为90℃。
进一步的,步骤(2)中,按照质量分数计,每85份蓝蛤肉酶解冻干粉、D-木糖和L-半胱氨酸混合物,加入1000份去离子水。
进一步的,所述风味蛋白酶为Flavourzyme风味蛋白酶;所述复合蛋白酶为Novozym复合蛋白酶11039;风味蛋白酶的添加量占蓝蛤肉重量的0.2%,复合蛋白酶的添加量占蓝蛤肉重量的0.2%。
进一步的,所述Novozym复合蛋白酶11039与Flavourzyme风味蛋白酶的质量比为1:1。
本发明的有益效果:
(1)、本发明以蓝蛤为原材料,选材天然、营养、健康。采用超高压耦合酶解与常压酶解相比酶解程度更高,产生了更多的低分子肽和游离氨基酸,超高压耦合酶解制备的蓝蛤肉酶解液风味抗氧化剂中游离氨基酸和5ˊ-核苷酸的含量显著下降,而有机酸的含量有所上升。根据电子舌结果可知,超高压耦合酶解制备的蓝蛤肉酶解液风味抗氧化剂的味觉特征与蓝蛤酶解液相比发生变化,表现为苦味、鲜味和涩味回味降低,而浓厚味增加,其味感更加协调。
(2)、本发明蓝蛤肉酶解液风味抗氧化剂能显著增强其ABTS自由基清除活性,由原来的1233.1μM Trolox/mL提升至2253μM Trolox/mL;蓝蛤肉酶解液风味抗氧化剂还原力由0.319提升至0.978;蓝蛤肉酶解液风味抗氧化剂对Fe2+螯合活性有显著提升;蓝蛤肉酶解液风味抗氧化剂对酶解液的羟自由基清除活性有显著的提升。因此蓝蛤肉酶解液风味抗氧化剂具有抗氧化性,可以在一定程度上避免油脂氧化,同时提高了调味料的营养性和功能性。
(3)、本发明采用喷雾干燥方式制备的具有抗氧化功能的耐高温蓝蛤调味基料具有低水分和高溶解度的特点,并且能有效降低蓝蛤肉酶解液风味抗氧化剂吸湿性;该耐高温蓝蛤调味基料固体混合物呈球状,无粘结现象并且表面光滑无凹陷和孔隙,并且内源荧光强度因添加而明显降低,这表明壳聚糖和阿拉伯胶埋效果良好,可以对氧气的渗透有隔离作用,保护核心材料免受氧化。
(4)、本发明的具有抗氧化功能的耐高温蓝蛤调味基料具有很好的热稳定性,壳聚糖和阿拉伯胶的加入该耐高温蓝蛤调味基料的挥发性风味物质有良好的保护效果,能有效延缓和减少核心材料损失,使其用于火锅锅底调味料时具有持香效果。
附图说明
图1本发明实施例1以及其它壁材的固体混合物的T2弛豫时间谱图;
图2本发明实施例1以及其它壁材的固体混合物的粒径分布(a)和平均粒径(b)图;
图3本发明实施例1以及其它壁材的固体混合物的Zeta电位图;
图4本发明实施例1与空白对照组的固体混合物的荧光光谱图;
图5本发明实施例1以及其它壁材的固体混合物的光学显微镜图(100×);
图6本发明实施例1以及其它壁材的固体混合物的扫描电镜图;
图7本发明实施例1以及其它壁材的固体混合物的热重分析图;
图8本发明实施例1以及其它壁材的固体混合物的鲜味释放分析图;
图9本发明实施例1、对比例1-3不同酶解液的DPPH自由基清除活性图;
图10本发明实施例1、对比例1-3不同酶解液的ABTS自由基清除活性图;
图11本发明实施例1、对比例1-3不同酶解液的还原力活性图;
图12本发明实施例1、对比例1-3不同酶解液的Fe2+螯合活性图;
图13本发明实施例1、对比例1-3不同酶解液的羟自由基清除活性图;
图14本发明实施例1超高压蓝蛤肉酶解冻干粉F1和对比例1常压蓝蛤肉酶解冻干粉F2中分子量的分布图;
图15本发明实施例1、对比例1-3不同酶解液的电子舌PCA分析图。
具体实施方式
实施例1
(1)超高压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤脱壳取肉,取1kg蓝蛤肉加入1kg去离子水后均质,添加2g风味蛋白酶Flavourzyme 500MG(500LAPU/g)和2g碱性蛋白酶Novozym 11039(1.2AU-A/g),用浓度为1.0mol/L的NaOH溶液调pH为7.0,在250Mpa条件下高压处理1h,后置于50℃水浴锅中酶解4h,酶解结束后100℃下灭活10min;在4℃下以5120×g离心15min,将上清液冷冻干燥后,得到超高压蓝蛤肉酶解冻干粉,记为F1;
(2)酶解冻干粉-木糖-半胱氨酸混合液的配制
以0.50g超高压蓝蛤肉酶解冻干粉、0.20g D-木糖和0.15g L-半胱氨酸为原料,加入去离子水10mL,调pH至7.0,得到酶解冻干粉-木糖-半胱氨酸混合液,记为F1BC;
(3)蓝蛤肉酶解物风味抗氧化剂的制备
将酶解冻干粉-木糖-半胱氨酸混合液置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂,记为MR PF1;
(4)蓝蛤肉酶解物风味抗氧化剂热稳定性处理
a.壳聚糖-阿拉伯胶-风味抗氧化剂乳液的制备
壳聚糖(C)和阿拉伯胶(GA)按照质量比9:1混合,加入蓝蛤肉酶解物风味抗氧化剂,壳聚糖和阿拉伯胶总加入质量占的蓝蛤肉酶解物风味抗氧化剂质量的5%,在室温下以2000r/min搅拌1h,制得壳聚糖-阿拉伯胶-风味抗氧化剂乳液;
b.喷雾干燥
将壳聚糖-阿拉伯胶-风味抗氧化剂乳液在9000r/mi n条件下均质10min,然后喷雾干燥,喷雾干燥条件为:进料流量9mL/min,进风温度为145℃,出风温度为90℃,得到目标产物具有抗氧化功能的耐高温蓝蛤调味基料固体混合物。
一、实施例1具有抗氧化功能的耐高温蓝蛤调味基料固体混合物(以下简称:固体混合物)物理性质分析
1、检测上述固体混合物水分含量、溶解度、吸湿性及体积密度,结果如表1所示:
(1)水分测定:采用GB5009.3-2016《食品安全国家标准食品中水分的测定》中的直接干燥法进行测定。
(2)溶解度测定:在室温下将0.250g固体混合物溶于25mL去离子水中,搅拌速度为100rpm/min,搅拌时间为30min。在(离心力1400×g,5min)条件下离心并取10mL上清液烘干(105℃),根据10mL上清液干燥后的质量来计算溶解度。
(3)吸湿性测定:将0.500g固体混合物(一式三份)放置在含饱和氯化钠溶液(75.3%相对湿度)的干燥器中7天。然后,根据样品吸收的水分质量计算吸湿性,计算结果以每100g固体所吸收的水分质量来表示。
(4)体积密度的测定:将粉末倒入带刻度的10mL量筒中,摇匀振实,填充至量筒刻度线5mL处,记录填充的固体混合物质量(m),以及填充体积(V),然后计算出体积密度(g/cm3)。
表1固体混合物物理特征
水分含量是影响固体混合物稳定性的主要因素之一,当水分含量高时会增强固体混合物在贮藏过程中的氧化速率和微生物活性。如表1所示,实施例1具有抗氧化功能的耐高温蓝蛤调味基料固体混合物的水分含量为4.96%,其中水分含量显著高于空白组(P<0.05),原因可能是蓝蛤肉酶解物风味抗氧化剂在壳聚糖+阿拉伯胶壁材包裹下,阻碍了水分的快速扩散和逃逸—在喷雾干燥过程中。此外,有研究发现当食物中的水分含量低于5%时,便具有良好的稳定性。因此,喷雾干燥能赋予蓝蛤肉酶解物风味抗氧化剂良好的稳定性。
吸湿性和溶解度是评价粉末或固体混合物的重要参数,当固体混合物吸湿性较高时,容易从周围环境中吸收水分而产生粘性和结块,造成不利影响,而溶解度的高低则影响固体混合物在水体系中的应用。由表1可知,空白对照组的溶解度92.62%,壳聚糖+阿拉伯胶C+GA组的溶解度略有降低为90.57%(P>0.05),该结果表明添加壳聚糖+阿拉伯胶壁材对固体混合物的溶解性无显著影响。此外,添加壳聚糖+阿拉伯胶壁材对固体混合物的吸湿性有显著降低作用(P<0.05),吸湿性由39.35g水/100g固体降低到26.86g水/100g固体范围内,这表明添加壳聚糖+阿拉伯胶壁材能改善其吸湿性,更有利于固体混合物的贮藏;固体混合物的体积密度是固体混合物粉末包装、运输和重组的重要性能,它取决于颗粒间空间的程度或体积的间隙度。由表1可知,实施例1固体混合物的体积密度在0.71g/cm3,与空白对照组几者间无显著差异。综上所述,添加壳聚糖+阿拉伯胶壁材有利于降低固体混合物的吸湿性,水分含量稍有提高,而对溶解度以及体积密度均无显著影响。
二、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物水分分布和组成分析
1、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物的制备
将实施例1制备的蓝蛤肉酶解物风味抗氧化剂,未添加壁材壳聚糖+阿拉伯胶(C+GA)的蓝蛤肉酶解物风味抗氧化剂按实施例1制备作为空白对照(B);采用以下壁材替代实施例1中的壳聚糖+阿拉伯胶(C+GA):壳聚糖(C)、麦芽糊精(M)、麦芽糊精+阿拉伯胶(M+GA,质量比9:1)、壳聚糖+麦芽糊精+阿拉伯胶(C+M+GA,质量比4.5:4.5:1),其它同实施例1,得到固体混合物。
2、水分分布和组成分析测定:采用低场核磁共振(low-field nuclear magneticresonance,LF-NMR)测定固体混合物中水分分布与组成的情况,将固体混合物装入核磁管中,填装量为核磁管1cm高度处,采用Carr-Purcell-Meiboom-Gill脉冲序列测定凝胶样品的横向弛豫时间T2。测定条件:SFI(MHz)=22,P90(μs)=14,SW(KHz)=200,TR(ms)=2000,NS=8,τ(μs)=200,Echocnt=4000。所得CPMG指数衰减曲线采用仪器自带的软件进行繁衍得到T2图谱。上述6组不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物检测结果如图1所示:
由图1可知,6组固体混合物中的T2弛豫时间均在0.1~1ms范围内,这部分水为性质稳定的结合水。这表明固体混合物中不含活泼的自由水或者含量极少,这与水分含量测定结果一致,证实了固体混合物具有良好的贮藏性。
三、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物粒度分析
1、粒度分析测定:以超纯水作为分散剂,将固体混合物配制成浓度为1mg/mL的溶液,测定温度为25℃,测定波长640nm,折射率为1.52,吸收率为0.1。
2、粒径和粒径分布是评价固体混合物复原体系稳定性和均匀性的重要参数,由于粒径较大的体系易发生聚集效应,从而导致体系稳定性降低。如图2(a)所示,空白组的粒径分布在100~1200nm范围内,粒径分布较宽,这表明形成了大小不一的聚集体和颗粒。其中M+GA组的粒径分布朝着增大的趋势变化,并且出现两个峰值,表明体系更加不稳定。这可能是由于喷雾干燥制备的固体混合物存在异质性,即固体混合物粒径大小不均匀。而C、M、C+GA(实施例1)以及C+M+GA组中粒径分布变得集中,前三组的粒径分布在200~1000nm内,而C+M+GA组的粒径为200~600nm范围内。整体来看,除M+GA外,C、M、C+GA(实施例1)以及C+M+GA组固体混合物的粒径大小均较空白对照组更加均匀。
图2(b)是固体混合物的平均粒径图,由图可知,空白组的平均粒径为523nm,添加壁材后平均粒径均呈显著增大的趋势(P<0.05)。其中C+GA(实施例1)、M+GA及C+M+GA组的平均粒径最大,在738~793nm范围内。但整体来看,这些组的粒径均较小,其水溶液体系稳定。
四、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物Zeta电位分析
1、Zeta电位测定:使用超纯水为分散剂,配制浓度为1mg/mL的溶液,测定温度为25℃。
2、Zeta电位作为指示溶液中粒子表面电荷的指标,能决定粒子的分散和聚集,是表征体系稳定性的重要指标。较高的粒子表面电荷能增强粒子间的静电排斥力,并克服范德华力和疏水引力,从而起到维持体系稳定性的作用。
图3是固体混合物的Zeta电位值,由图可知,空白组的Zeta电位值最大,为-35.6mv。添加壁材后,其电位值均呈降低趋势(P<0.05),其中C、M、C+GA、M+GA、C+M+GA组的电位值分别为-31.6mv、-30.5mv、-28.6mv、-16.9mv、-20.9mv。上述结果表明壁材对固体混合物溶液体系的稳定性有降低作用,其中M+GA组的稳定性最差,该结果与粒径测定结果一致。
五、本发明实施例1及空白对照组内源荧光分析
1、内源荧光测定:用超纯水将固体混合物配制成浓度为0.5mg/mL的溶液,然后在该浓度下进行荧光扫描。测试条件:灵敏度:5;扫描速度:高速;EX缝宽:5nm;EM缝宽:5nm;当前波长EX:295nm;EM范围:320nm~450nm。
2、酪氨酸(tyrosine,Tyr)残基荧光分析常用于分析蛋白质结构变化。图4是固体混合物的内源荧光光谱图,由图可知,添加壳聚糖-阿拉伯胶壁材后,固体混合物溶液的荧光强度出现了降低,这表明加入壁材对Trp残基的暴露有掩蔽作用,原因是壁材将蓝蛤肉酶解液风味抗氧化剂包埋,导致其中的Trp残基不能完全暴露在周围溶剂中。
六、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物形态特征分析
1、显微镜观察
取固体混合物溶于去离子水(1:10,w:v)中溶解1min,后取约10μL溶液于载玻片中央,用显微镜进行观察(100×),图像是通过连接到显微镜上的照相机拍摄并分析。
2、显微镜观察结果
取固体混合物溶于去离子水(1:10,w:v)中溶解1min,后取约10μL溶液于载玻片中央,用显微镜进行观察(100×),图像是通过连接到显微镜上的照相机拍摄并分析。
3、不同的壁材会对固体混合物的包埋效果、疏水性造成影响
图5是固体混合物的光学显微镜图(100×)。由图可知,B组中未见球状物,这是由于固体混合物快速溶于介质中,分散为微小颗粒。而添加壁材的C组、M组、C+GA组、M+GA组、C+M+GA组中有明显的球状物存在,这是由于壁材具有一定的疏水性,在固体混合物-介质界面处因疏水相互作用而形成膜,从而延缓了介质的渗入和固体混合物的流出。并且M+GA组的固体混合物的表观粒径大小不一,差异最为明显。该结果与粒径和Zeta电位测定结果一致,即未添加壁材的固体混合物溶解迅速,形成了粒径较小的颗粒和稳定性高的体系,而添加壁材组的固体混合物溶解缓慢,粒径较大。
4、SEM结果
扫描电镜主要研究固体混合物的外部形貌,可用于确定固体混合物的包埋效果和稳定性。图6是固体混合物的SEM图,由图可知,固体混合物形态基本呈圆球形或不规则球形,这是由于在干燥过程中固体混合物外表面发生硬化收缩,而内部液滴气泡膨胀所致,B组、C组和C+GA组的固体混合物呈规则的圆球状并且表面光滑无明显的凹陷和褶皱,其中B组的固体混合物存在轻微的团聚粘黏现象,这可能是由于在干燥过程中发生浓缩而黏度增大所致。M组、M+GA组和C+M+GA组的固体混合物呈不规则球形,并且外表存在明显的凹陷、褶皱和孔隙,凹陷和褶皱是由于固体混合物乳液在喷雾干燥过程中,液滴表面水分快速蒸发,固体混合物膜壁瞬间收缩引起的。而孔隙的出现则是在喷雾干燥时,高热量使固体混合物表面水分蒸发的速率远大于内部水分扩散迁移至表面的速率,导致固体混合物内外产生压力差,内部压力大,液滴在干燥时内部膨胀所致。其中C+M+GA组发生严重的粘黏现象,影响着固体混合物的分散性和其他性质。综上所述,C组和C+GA组的固体混合物表观结构最佳,外形圆整,无凹陷和孔隙,这从稳定性的角度来看,它对氧气的渗透有隔离作用,可以很好地保护核心材料免受氧化。
七、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物热重分析
1、热重分析:称取4mg固体混合物加入铝坩埚中,氮气流量为20cm3/min,以10℃/min的速率从30℃上升至600℃,进行热重分析(Thermogravimetric analysis,TGA)
2、热重分析结果:图7是固体混合物的热重分析结果,其中50~150℃为第一反应阶段,在该范围内各组固体混合物的损失较小,约为6%左右,这部分损失归因于自由水的蒸发和结合水在高温下转变为自由水而蒸发;随着温度的升高,固体混合物的损失速率明显加快,其中150~500℃为第二反应阶段。在该阶段,B组的损失最大,在500℃时损失了70%左右,C组、M组、M+GA组、C+M+GA组的固体混合物的损失率相似,约为60%,而C+GA组的损失率最小,约为55%。该阶段的质量损失可能是壁材热降解所致;当温度继续升高至550℃时,各组固体混合物的热损失率为最大,从图中可见陡峭的下降曲线,此时B组固体混合物损失了90%,而M组、C+M+GA组、C组、M+GA组的质量损失在60~70%范围内,C+GA组的质量损失为55%。综上所述,添加壁材后固体混合物的热稳定性明显提高,其中实施例1添加壳聚糖+阿拉伯胶的稳定性最好。
八、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物鲜味释放率测定
1、鲜味释放率测定:准确称取0.500g固体混合物装入滤袋中,将其放置于100mL去离子水中(30℃)浸泡不同时间(0、0.5、1、2、4、8、16、32、60min),分别取不同浸泡时间的固体混合物溶液进行电子舌的测定。鲜味释放率计算方式如下:
UR(%)=Ux/U60min×100%
Ux:各种固体混合物不同浸泡时间溶液的鲜味强度值;
U60min:各种固体混合物浸泡60min溶液的鲜味强度值;
2、鲜味释放率分析
鲜味物质在烹饪过程中会高温分解,从而导致鲜味强度减弱,采用固体混合物技术将其进行包埋并达到缓慢释放的效果便可能有效减少鲜味减弱等问题。图5-图8是不同固体混合物在40℃水中的鲜味释放率分析图,由图可知,B组的鲜味释放速率最快,鲜味在0.5min时释放了72.96%,而4min时便已经释放了91.51%,在32min时基本完全释放(98.26%);添加壁材后固体混合物的鲜味释放速率延缓,其中C组和M组的缓释效果较差,在0.5min时释放率为44.44~55.31%,4min时释放了85.50~86.39%,由此可见,添加壳聚糖或麦芽糊精只能短暂延缓其释放,当固体混合物在水中浸泡2min时外壁便快速溶解而释放出芯材;M+GA组和C+M+GA组的缓释效果较好,在0.5min时释放率为28.03~30.80%,4min时仅释放了50%左右,直到32min时才释放了90%左右;而C+GA组的鲜味缓释效果最佳,其在0.5min时释放了13.95%,4min时才释放了38.08%,32min时释放了80%。产生良好的缓释效果是由于添加的C+GA将蓝蛤肉酶解液风味抗氧化剂芯材包裹于其中并形成密闭性良好的固体混合物,而C+GA壁材的水溶解速率较慢,从而起到阻碍水分进入的屏障效应。
九、不同壁材的蓝蛤风味抗氧化剂乳液制备具有抗氧化功能的耐高温蓝蛤调味基料固体混合物挥发性化合物测定
由表2和表3可知,共检测出醛类、酮类、芳香族化合物、酯类、酸类、酚类、烃类、呋喃类、含硫化合物、含氮化合物。醛类物质包括异戊醛、2-甲基丁醛、苯甲醛和苯乙醛,主要呈水果、坚果味,由表可知,与B组相比,添加壁材组的固体混合物中醛类种类和含量均更高,其中C+GA组的最高;2-戊酮和2-壬酮是主要的呈味酮类,表现为水果及脂肪香,但由于阈值较高,在较低含量时对固体混合物的风味构成贡献较小;甲苯和二丁基羟基甲苯是检测出的两种芳香族化合物,它们对风味贡献极小,其中二丁基羟基甲苯是一种商业常用的抗氧化剂,可能为固体混合物的抗氧化作出贡献;酸类、酚类和酯类的阈值较高,因此对风味无贡献;呋喃类是构成样品肉味风味的重要风味物质,其中包括糠醛、2-甲基-3-呋喃硫醇、糠(基)硫醇、2-正戊基呋喃和双(2-甲基-3-呋喃基)二硫,风味特征现出咖啡和烤肉香,由表可知,B组中的呋喃类物质含量和种类均最低(3种,7.56%),添加壁材能明显减少其损失,其中C+GA组的呋喃含量最高(28.21%);含硫化合物的存在对固体混合物风味的影响有好有坏,其中二甲基二硫、3-甲基噻吩和2-甲基噻吩呈刺鼻的硫磺味,贡献出不好的风味。而二氢-2-甲基-3(2H)-噻吩酮和1-(2-呋喃基甲基)-1H-吡咯贡献出焙烤香味和咖啡香味;含氮化合物中对风味贡献较大的是2-乙基-3-甲基吡嗪和2-乙基-5-甲基吡嗪等吡嗪类物质,能贡献出巧克力和坚果风味。
由上述可知,对固体混合物风味贡献较大的物质为醛类、呋喃类、含硫化合物和含氮化合物,其中呋喃类对固体混合物的烤肉风味贡献最大。在喷雾干燥过程中,固体混合物风味物质有所损失,添加壁材能有效延缓和减少其损失-由于形成良好的包埋物,其中C+GA组的保护效果最佳。
表2不同壁材的固体混合物中挥发性成分的数量及相对含量
表3不同壁材的固体混合物中的挥发性化合物分析
ND:未检出;a GC-MS鉴定;b正构烷烃C8~C20在HP-5MS柱上测定保留指数;c气味描述主要来自以下在线数据库(http://www.odour.org.uk/,http://www.flavornet.org/,https://www.chemicalbook.com/ProductIndex.aspx).d RI:保留指数,MS:质谱库。
综上所述,喷雾干燥制备的固体混合物具有低水分和高溶解度的特点,并且添加壁材后能有效降低其吸湿性,但是会导致固体混合物水溶液的粒径增大和Zeta电位降低,使得溶液稳定性有所降低;分析形态特征可知,固体混合物呈球状,其中添加壳聚糖组和添加壳聚糖+阿拉伯胶组的效果最佳,无粘结现象并且表面光滑无凹陷和孔隙,并且内源荧光强度因添加壁材而明显降低,这表明其包埋效果良好;热重分析表明,添加壁材后固体混合物的热稳定性明显提高,其中添加壳聚糖+阿拉伯胶的效果最佳;鲜味释放结果表明,空白组的鲜味释放速率最快,4min时便已经释放了91.51%,而壳聚糖+阿拉伯胶组浸泡32min时才释放了80%;GC-MS结果表明,壳聚糖+阿拉伯胶的加入对固体混合物的挥发性风味物质有良好的保护效果。综上所述,添加壳聚糖+阿拉伯胶壁材可以很好的提高对固体混合物的稳定性、鲜味缓释和风味保留效果。
对比例1
(1)常压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤脱壳取肉,取1kg蓝蛤肉加入1kg去离子水后均质,添加2g风味蛋白酶Flavourzyme 500MG(500LAPU/g)和2g碱性蛋白酶Novozym 11039(1.2AU-A/g),用浓度为1.0mol/L的NaOH溶液调pH为7.0,置于50℃水浴锅中常压酶解4h,酶解结束后100℃下灭活10min;在4℃下以5120×g离心15min,将上清液冷冻干燥后,得到常压酶解冻干粉,记为F2;
(2)酶解冻干粉-木糖-半胱氨酸混合液的配制
以0.50g常压酶解冻干粉、0.20g D-木糖和0.15g L-半胱氨酸为原料,加入去离子水10mL,将反应体系pH调为7.0,得到酶解冻干粉-木糖-半胱氨酸混合液,记为F2BC;
(3)蓝蛤肉酶解物风味抗氧化剂的制备
将酶解冻干粉-木糖-半胱氨酸混合液置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂,记为MR PF2;
对比例2未添加D-木糖和L-半胱氨酸的热反应产物阴性对照1
(1)超高压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤脱壳取肉,取1kg蓝蛤肉加入1kg去离子水后均质,添加2g风味蛋白酶Flavourzyme500MG(500LAPU/g)和2g碱性蛋白酶Novozym 11039(1.2AU-A/g),用浓度为1.0mol/L的NaOH溶液调pH为7.0,在250Mpa条件下高压处理1h,后置于50℃水浴锅中酶解4h,酶解结束后100℃下灭活10min;在4℃下以5120×g离心15min,将上清液冷冻干燥后,得到超高压蓝蛤肉酶解冻干粉,记为F1;
(2)酶解冻干粉-木糖-半胱氨酸混合液的配制
以0.50g超高压蓝蛤肉酶解冻干粉加入去离子水10mL,将反应体系pH调为7.0;置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂,记为F1 NC;
对比例3未添加D-木糖和L-半胱氨酸的热反应产物阴性对照2
(1)常压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤脱壳取肉,取1kg蓝蛤肉加入1kg去离子水后均质,添加2g风味蛋白酶Flavourzyme 500MG(500LAPU/g)和2g碱性蛋白酶Novozym 11039(1.2AU-A/g),用浓度为1.0mol/L的NaOH溶液调pH为7.0,置于50℃水浴锅中常压酶解4h,酶解结束后100℃下灭活10min;在4℃下以5120×g离心15min,将上清液冷冻干燥后,得到常压酶解冻干粉,记为F2;
(2)以0.50g常压酶解冻干粉为原料,加入去离子水10mL,将反应体系pH调为7.0;置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂,记为F2 NC;
十、实施例1、对比例1-3分别将未添加D-木糖和L-半胱氨酸的热反应产物作为阴性对照(F1NC和F2NC),未加热反应作为空白对照(F1BC和F2BC)。性能检测如下:
1、检测方式
(1)、DPPH自由基清除活性测定:移取10mL样液和10mL DPPH溶液(0.2mmol/L,溶于无水乙醇),混合均匀后,于室温下避光放置30min,随后在517nm处测定其吸光度(A2);将10mL样液和10mL无水乙醇混合,其它操作同上,记为A1;将10mL DPPH溶液和10mL无水乙醇混合,其它操作同上,记为A0;按照下式计算DPPH自由基清除率。
R=(1-(A2-A1)/A0)×100%
(2)、ABTS自由基清除活性测定:将配制好的ABTS溶液(7mmol/L)与过硫酸钾溶液(2.4mmol/L)按1:1的比例混合后30℃避光孵育12h,制成ABTS自由基储备溶液。用前以pH7.4的磷酸缓冲溶液(0.2mol/L)为溶剂稀释ABTS自由基储备溶液至734nm下吸光度为0.7±0.02,取10μL样品溶液和4mL的ABTS工作液,振荡混匀,静置6min后在734nm处测定吸光度。同时以6-羟基-2,5,7,8-四甲基色烷-2-羧酸(Troloxμmol)做标曲,依据方程y=251.96x+41.516,R2=0.987计算Trolox当量。
(3)、还原力测定:将0.5mL样品溶液与2.5mL磷酸钠缓冲液(pH6.6,0.2mol/L)和2.5mL铁氰化钾(1g/mL,K3{Fe(CN)6})混合,后于50℃水浴20min。在室温下加入2.5mL的三氯乙酸(TCA,10g/mL),在3000×g下离心15min,收集上清液。取1mL上清液加入超纯水10mL和1mL FeCl3(0.1g/mL),充分振荡后静置10min。使用紫外-分光光度计在700nm处读取混合物的吸光度。吸光度越高则还原力越高。用吸光度表示结果。
(4)、Fe2+螯合活性测定:取0.2mL样品溶液与0.1mL FeCl2(2mmol/L)、4mL超纯水室温下混匀,静置5min后加入0.2mL菲啰嗪(5mmol/L)。室温下混匀5min后以2000rpm离心10min,取上清液在562nm处测定吸光度(As)。取超纯水代替样品溶液按上述条件测定吸光度(A0),计算公式如下:
Fe2+螯合率(%)=(A0-AS)/A0×100%
(5)、羟自由基清除活性测定:取0.1mL样品溶液,加入2mL的FeSO4(9mmol/L)、2mL的水杨酸-乙醇溶液(9mmol/L)和2mL的H2O2溶液(8.8mmol/L),混匀后于37℃水浴30min,在510nm处下测得A1;用去离子水代替样品如上述方法测得A。用去离子水代替水杨酸-乙醇溶液测得A0,计算清除率公式如下:
-OH清除率(%)=(1-(A1-A0)/A)×100
2、结果与分析
(1)DPPH清除活性分析
DPPH自由基可通过氢的贡献被抗氧化物质清除,目前DPPH自由基清除活性测定已被用作评价物质抗氧化能力的标准。对超高压组和常压组的DPPH自由基清除活性分别进行方差分析,结果如图9所示,F1 BC、F1NC、MR PF1的DPPH自由基清除活性分别为67.8%、67.2%、83.4%,F2 BC、F2 NC、MR PF2的DPPH自由基清除活性分别为74.3%、75.1%、81.2%。与BC组相比,MR PF组的DPPH自由基清除率从67.8%~75.8%增加到81.2~83.4%(P<0.05),其原因可能是MR PF组反应中产生的黑色素和低分子量的杂环化合物导致的。
超高压酶解冻干粉F1的分子量低于常压酶解冻干粉F2,由图9可知,MR PF1的DPPH自由基清除活性优于MR PF2。与BC组相比,NC组的DPPH自由基清除活性差异不显著(P>0.05),表明只加热对DPPH自由基清除活性的变化无明显影响。综上所述,MR PF组反应后能增强蓝蛤酶解液的DPPH自由基清除活性,且超高压酶解液的MR PF1反应产物的DPPH自由基清除活性优于常压酶解液的MR PF2反应产物。
(2)ABTS自由基清除活性分析
ABTS自由基清除法是常见的用于评价抗氧化能力的一种方法,其原理是ABTS二铵盐经过硫酸钾氧化后生成稳定的ABTS自由基,该物质呈蓝绿色,当加入具有抗氧化效果的物质后吸光度会降低。
从图10可看出,蓝蛤酶解液自身就具有一定的清除ABTS自由基能力(1233.1±55.1μM Trolox/mL),这是由于酶解过程中,蛋白质被水解成具有抗氧化能力的多肽和氨基酸,使其清除自由基的能力得到增强。组内差异性分析表明,与酶解液相比,热反应产物(F1NC:1643.3μMTrolox/mL vs F2NC:1636.7μMTrolox/mL)及MR PF组反应产物(MR PF1:2253μMTrolox/mL vs MR PF2:2226.7μMTrolox/mL)具有更高的清除活性。超高压MR PF1产物的ABTS自由基能力优于常压MR PF2产物。
(3)还原力活性分析
由图11可知,酶解液自身同样具有一定的还原力,但还原力较低(F1BC:0.319vsF2BC:0.195),热反应后,其对应的加热产物的还原力有所增长但增幅小(F1NC:0.361vsF2NC:0.299)。超高压耦合酶解液及常压酶解液的MRPF组还原力(MR PF1:0.978vs MR PF2:0.937)显著增强(P<0.05)。超高压处理组的还原力更高,原因可能是产生了更多的游离氨基酸,进而提升其还原力。
(4)Fe2+螯合活性分析
如图12所示,各组对Fe2+均有一定的螯合能力,但整体螯合能力较弱,其中BC组的螯合能力最低(F1BC:21.01%vs F2BC:19.59%),其对应的热反应产物的螯合能力有所增强,而MR PF组反应产物的Fe2+螯合活性(MR PF1:33.56%vs MR PF2:30.09%)显著提升(P<0.05),超高压处理组比常压处理组螯合能力更强。而Fe2+金属离子对人体中部分酶的活性有所影响,与Fe2+螯合形成稳定的螯合物,减少自由基链式反应的进行,表现出一定的抗氧化作用。
(5)羟自由基清除活性分析
如图13所示,各组样品对羟自由基均有良好的清除效果,对照组、阴性对照组及MRPF2反应产物各组差异显著(P<0.05),其中空白组的清除效果最低(F1BC:77.14%vs F2BC:74.61%),阴性对照组产物清除效果次之(F1NC:80.88%vs F2NC:79.39%),而MR PF反应产物则最高(MRPF1:85.72%vs MRPF2:83.80%)。上述结果表明酶解液自身就有较高的羟自由基清除活性,加热对酶解液的羟自由基清除活性也有显著的提升,其中MRPF反应产物提升最为显著,超高压处理对其也有一定的增强。
综上所示,本发明实施例1和对比例1蓝蛤超高压和常压酶解液MR PF1反应产物DPPH自由基清除活性、ABTS自由基清除活性、还原力、Fe2+螯合活性和羟自由基清除活性显著提升(P<0.05),且超高压MR PF1的抗氧化能力均优于常压的MR PF2)。热反应产物(阴性对照组)与酶解液相比,除羟自由基清除活性外,各抗氧化指标间差异不显著(P>0.05)。
综上所述,本发明实施例1的蓝蛤肉酶解物风味抗氧化剂(MR PF1)可以作为一种良好的抗氧化剂使用。
十一、实施例1、对比例1的F1和F2中分子量分布的测定
1、检测:采用凝胶色谱法测定酶解冻干粉中肽分子量的分布。流动相由70%(v/v)乙腈和30%(v/v)超纯水组成;流速0.5mL/min,柱温30℃,检测波长为220nm,样品进样量10μL。所用标准品如下:细胞色素C(12500Da)、抑酚酶(6500Da)、杆菌酶(1450Da)、乙氨酸-乙氨酸-酪氨酸-精氨酸(451Da)、乙氨酸-乙氨酸-乙氨酸(189Da)。肽分子量分布划分为<180Da、180-500Da、500-1000Da、1000-2000Da、2000-3000Da、3000-5000Da和>5000Da。
2、结果:图14显示了F1和F2中肽分子量的分布情况。在两种酶解冻干粉中,分子量<180Da和180-500Da的肽含量占比最高,其中F1中分子量<180Da和180-500Da的肽分别占64.12%,29.49%,而在F2中分别占36.60%,53.90%。该结果表明,超高压耦合酶解程度更高,产生了更多的低分子肽。
十二、实施例1、对比例1-3的酶解液游离氨基酸的测定
1、检测:用于游离氨基酸的测定。将样品与氨基水杨酸以4:1(v:v)的比例在离心管中混合,然后将混合物置于4℃下使蛋白质沉淀1h。离心条件为4℃下以11,963×g离心15min,上清液过0.22μm滤膜,最后使用自动氨基酸分析仪进行测定和分析。重复3次。
2、结果:游离氨基酸(free amino acid,FAAs)对味觉和挥发性风味化合物的形成均起着重要的作用。如表4所示,在所有样品均检测到15种FAAs,根据呈味特性可分为不同类别:鲜味、甜味以及苦味氨基酸。
从表4可知,无论哪种类型的氨基酸,其在F1BC中的含量总是高于F2BC,其中F1BC的总游离氨基酸高达1432mg/100g,而F2BC的是1300mg/100g。F1BC的苦味氨基酸含量占比为47.90%,甜味氨基酸占31.56%,鲜味氨基酸占20.53%。此外,F2BC中苦味氨基酸含量占比为51.15%,甜味氨基酸占30.92%,鲜味氨基酸占17.92%。这表明酶解方式对FAAs的含量和不同滋味氨基酸的比例有显著的影响(P<0.05),超高压耦合酶解不但提高了FAAs的含量,还降低了苦味氨基酸的比例。
MRPF组所有氨基酸含量均显著降低(P<0.05),其中总游离氨基酸的含量减少了39.66~52.69%,鲜味氨基酸减少了51.70~53.6%,甜味氨基酸减少了46.24~53.98%,苦味氨基酸了30.17~50.98%。
不同氨基酸的味觉阈值是不同的,因此不能只从氨基酸含量的角度来分析贡献度大小。为了评价化合物对味觉的影响,味觉活性值(taste activity value,TAV)是一个非常有用的指标,TAV是根据样品的含量与它在水或其他简单基质中测定的阈值之间的比值计算的,TAV大于1的化合物被认为对食品滋味有显著的贡献。
如表4所示,谷氨酸(Glu)、丙氨酸(Ala)、赖氨酸(Lys)、蛋氨酸(Met)、缬氨酸(Val)、精氨酸(Arg)和组氨酸(His)的TAV大于1。其中谷氨酸(Glu)的TAV最高,对样品的滋味贡献度最大。丙氨酸(Ala)具有令人愉快的甜味,在海鲜中的含量较高,而其他的氨基酸也会贡献出明显的苦味。
表4实施例1、对比例1-3的酶解液中游离氨基酸的变化
注:Glu,谷氨酸;Asp,天冬氨酸;Gly,甘氨酸;Ala,丙氨酸;Ser,丝氨酸;Thr,苏氨酸;Pro,脯氨酸;Lys,赖氨酸;Met,蛋氨酸;Val,缬氨酸;Arg,精氨酸;Ile,异亮氨酸;Leu,亮氨酸;His,组氨酸;Phe,苯丙氨酸。*必需氨基酸。
十三、实施例1、对比例1-3的酶解液有机酸的测定
1、有机酸检测:
(1)标准曲线:混合标准溶液中有4种有机酸:包括苹果酸、乳酸、柠檬酸、琥珀酸配制成一系列浓度标准溶液,0.01、0.02、0.04、0.06、0.08、0.10mg/mL,将6个浓度梯度混合标准溶液在相同的液相条件下进样,在将得到每种有机酸的峰面积对浓度(mg/mL)作图绘制4种有机酸标准曲线,并采用外标法进行定量。所得标准曲线依次为:y=719.710x+0.527(R2=0.999),y=535.490x-0.1433(R2=0.998),y=903.410x-0.0369(R2=0.999),y=634.550x-0.614(R2=0.998)。
(2)样品前处理:取5mL样品加入0.25mL高氯酸(5%,v∶v)匀浆静置20min,离心(6150×g,10min,4℃),取上清液过0.45μm滤膜,待高效液相色谱进样分析。
(3)色谱条件:C18柱(5μm,4.6mm×250mm);柱温:25℃;检测器:紫外吸收检测器,检测波长:205nm;流动相:甲醇∶磷酸二氢钾(0.01mol/L,pH=2.8)=5∶95(v∶v);流速:0.5mL/min;进样量5μL;等度洗脱。重复3次。
2、结果分析:如表5所示,在所有样品中鉴定出了4种有机酸,包括苹果酸、乳酸、柠檬酸和琥珀酸,这些有机酸不仅具有酸味,而且还能提供鲜味。琥珀酸是一些海鲜特别是贝类的主要味觉活性成分之一,琥珀酸及其钠盐可贡献特殊的酸味和鲜味。琥珀酸是所有样品中的最主要的有机酸,占总有机酸含量的47.87%~79.94%,含量在21.73~32.82mg/mL范围内,TAV均大于200。因此琥珀酸对蓝蛤酶解液及其固体混合物的鲜味贡献起着极其重要的作用。与F2相比,在F1中琥珀酸含量更高(P<0.05),表明超高压耦合酶解可产生更多的琥珀酸,原因可能是琥珀酸盐内部的连接在高压下断裂从而释放出游离琥珀酸。此外,MRPF组琥珀酸的含量略有减少(P>0.05)。
表5实施例1、对比例1-3的酶解液中有机酸的变化
在所有样品中,乳酸是含量仅次于琥珀酸的有机酸,占总有机酸的15.96~34.66%,含量在6.34~15.73mg/mL,TAV均大于5。乳酸是发酵食品中一种重要的滋味物质,也是水产品中主要的有机酸。在所有样品中柠檬酸的含量均明显低于乳酸的含量,但柠檬酸的阈值较低,对味觉贡献度与乳酸的相近。与上述三种有机酸相比,苹果酸的滋味贡献最少。总体来看,超高压处理对有机酸的释放有促进效果,并且MRPF组有机酸总含量有所增加。
十四、实施例1、对比例1-3的酶解液的5′-呈味核苷酸的测定
1、检测:
(1)标准曲线:混合标准溶液中有3种核苷酸:包括5′-AMP、5′-IMP、5′-GMP配制成一系列浓度标准溶液,0.01、0.02、0.04、0.06、0.08、0.10mg/mL。所得标准曲线依次为:y=43107x+8.0529(R2=0.993),y=33256x+12.0930(R2=0.992),y=47089x-9.7841(R2=0.993)。
(2)样品前处理:取5mL样品加入15mL高氯酸(5%,v:v),匀浆后与在4℃条件下以4500×g离心10min,取上清液。沉淀中加入10mL高氯酸(5%,v:v),匀浆离心(条件同上)。合并两次上清液,用5mol/L KOH调pH至6.75,过0.22μm滤膜。
(3)色谱条件:C18柱(5μm,4.6mm×250mm);流动相:pH为4.5的0.05mol/L的KH2PO4-甲醇溶液,临用前超声脱气;流速:0.8mL/min;进样量:20μL;紫外检测器波长:254nm;采用梯度洗脱,程序如下:0-10min:97.5%KH2PO4:2.5%甲醇(97.5%:2.5%);10-17min:85%KH2PO4:15%甲醇;17-20min:97.5%KH2PO4:2.5%甲醇。重复3次。
2、检测结果:
鲜味被定义为谷氨酸引起的特征味觉,包括5ˊ-GMP、5ˊ-IMP和5ˊ-AMP在内的5ˊ-核苷酸也能提供一定的鲜味。此外,还发现5ˊ-核苷酸和鲜味氨基酸之间存在协同作用,当它们以一定的比例混合时能产生更强烈的鲜味。如表6所示,在所有样品中均检测出5ˊ-GMP和5ˊ-AMP,但5ˊ-IMP的浓度过低,未检出。GMP主要由RNA的破坏性分解产生,而IMP和AMP则由动物死亡后ATP降解而产生,这一过程归因于肌肉内源性酶。ATP分解有两条途径,分别为ATP→ADP→AMP→IMP→HxR→Hx和ATP→ADP→AMP→AdR→HxR→Hx。因此,推测蓝蛤中ATP的分解途径为第二条途径。表6显示了5ˊ-核苷酸的含量和TAV,其中最主要的核苷酸是5ˊ-GMP,其次是5ˊ-AMP。5ˊ-GMP在F1BC、F1NC和MRPF1中的含量及TAV分别为32.97mg/100mL(TAV,2.64),30.62mg/100mL(TAV,2.45)和26.52mg/100mL(TAV,2.12);而在F2BC、F2NC和MRPF2中的含量及TAV分别为30.96mg/100mL(TAV,2.48),28.11mg/100mL(TAV,2.25)和23.38mg/100mL(TAV,1.87)。由此可知5ˊ-GMP对蓝蛤酶解液以及MRPF1组的滋味均有贡献,由于TAV大于1。此外,MRPF1组5ˊ-GMP略有下降(P<0.05),原因可能是高温分解。与5ˊ-GMP相比,5ˊ-AMP的TAV小于1,因此对滋味的影响可忽略不计。
表6实施例1、对比例1-3的酶解液中5ˊ-核苷酸的变化
注:15′-GMP,5′-鸟苷酸;5′-AMP,5′-腺苷酸。
十五、电子舌的测定
1、检测:取待测样品4mL加去离子水定容至100mL,倒入电子舌专用杯中,样品体积约为70mL,然后用电子舌在室温下检测各样品的酸味、苦味、苦味回味、涩味、涩味回味、咸味、鲜味和浓厚味。每个样品循环4次,结果取后三次测定结果。测试前已对电子舌设备进行自检和校准。
2、检测结果:表7是样品稀释25倍(v:v)后测定的电子舌结果。由表7可知,所有样品中均未检出酸味、涩味、苦味回味、咸味(响应值分别小于-20、-1、0和-7,数值低于0则表示人类味觉不能感知),而苦味和鲜味的响应值明显高于其他滋味。与F2相比,F1具有较高的浓厚味和鲜味响应值,并且苦味和涩味回味较低(P<0.05)。而MRPF1组、MRPF2组的苦味、鲜味、涩味回味均显著降低(P<0.05);相反,浓厚味的响应值显著增加(P<0.05)。图15对不同传感器得到的响应值进行了PCA分析,PC1和PC2的贡献率分别为50.99%和41.10%,累积贡献率为92.09%。因此,PC1和PC2包含了大量信息,可以反映样本的总体特性。图中所有样品的数据采集点分散,现象表明,MRPF1组、MRPF2组的味道特征发生了变化。
表7实施例1、对比例1-3的酶解液的电子舌分析
综上所述,与对比例1常压酶解相比,实施例1超高压耦合酶解程度更高,产生了更多的低分子肽和FAAs;实施例1蓝蛤肉酶解物风味抗氧化剂中FAAs和5ˊ-核苷酸的含量显著下降,而有机酸的含量有所上升。根据电子舌结果可知,MRPF1组(本发明实施例1)的味觉特征发生变化,表现为苦味、鲜味和涩味回味降低,而浓厚味增加,其味感更加协调;此外,电子鼻和GC-MS结果表明,MRPF1具有腥味特征的醛类明显降低,具有肉香味的呋喃和含硫化物含量增加。
以上仅为本发明的具体实施例而已,并不用于限制本发明,对于本领域的技术人员来说,本发明可以有各种更改和变化。凡在本发明的精神和原则之内,所作的任何修改、等同替换、改进等,均应包含在本发明的保护范围之内。
Claims (5)
1.一种持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其特征是:
具体步骤如下:
(1)超高压酶解制备蓝蛤肉酶解冻干粉
将蓝蛤肉用风味蛋白酶和复合蛋白酶酶解,在250Mpa条件下高压处理1h后,进行酶解,制得蓝蛤肉酶解冻干粉;
(2)酶解冻干粉-木糖-半胱氨酸混合液的配制
以蓝蛤肉酶解冻干粉、D-木糖和L-半胱氨酸为原料,按照质量比5:2:1.5混合,搅拌均匀后,加入去离子水,调pH值至7.0,得到酶解冻干粉-木糖-半胱氨酸混合液;
(3)蓝蛤肉酶解物风味抗氧化剂的制备
将酶解冻干粉-木糖-半胱氨酸混合液置于油浴具塞容器中,密封,在120℃下油浴反应1.5h,得到蓝蛤肉酶解物风味抗氧化剂;
(4)蓝蛤肉酶解物风味抗氧化剂热稳定性处理
a.壳聚糖-阿拉伯胶-风味抗氧化剂乳液的制备
壳聚糖和阿拉伯胶按照质量比9:1混合,加入蓝蛤肉酶解物风味抗氧化剂,其中,壳聚糖和阿拉伯胶总加入质量占的蓝蛤肉酶解物风味抗氧化剂质量的5%,在室温下以2000r/min搅拌1h,制得壳聚糖-阿拉伯胶-风味抗氧化剂乳液;
b.喷雾干燥
将壳聚糖-阿拉伯胶-风味抗氧化剂乳液在9000r/min条件下均质10min,然后喷雾干燥,得到具有抗氧化功能的耐高温蓝蛤调味基料。
2.根据权利要求1所述的持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其特征是:喷雾干燥条件为:进料流量9mL/min,进风温度为145℃,出风温度为90℃。
3.根据权利要求1所述的持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其特征是:步骤(2)中,按照质量分数计,每85份蓝蛤肉酶解冻干粉、D-木糖和L-半胱氨酸混合物,加入1000份去离子水。
4.根据权利要求1所述的持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其特征是:所述风味蛋白酶为Flavourzyme风味蛋白酶;所述复合蛋白酶为Novozym复合蛋白酶11039;风味蛋白酶的添加量占蓝蛤肉重量的0.2%,复合蛋白酶的添加量占蓝蛤肉重量的0.2%。
5.根据权利要求1所述的持香型具有抗氧化功能的耐高温蓝蛤调味基料的制备方法,其特征是:所述Novozym复合蛋白酶11039与Flavourzyme风味蛋白酶的质量比为1:1。
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