CN108813527A - 一种含虾青素的调味品及其制备方法 - Google Patents
一种含虾青素的调味品及其制备方法 Download PDFInfo
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- JEBFVOLFMLUKLF-IFPLVEIFSA-N Astaxanthin Natural products CC(=C/C=C/C(=C/C=C/C1=C(C)C(=O)C(O)CC1(C)C)/C)C=CC=C(/C)C=CC=C(/C)C=CC2=C(C)C(=O)C(O)CC2(C)C JEBFVOLFMLUKLF-IFPLVEIFSA-N 0.000 title claims abstract description 114
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- MQZIGYBFDRPAKN-ZWAPEEGVSA-N astaxanthin Chemical compound C([C@H](O)C(=O)C=1C)C(C)(C)C=1/C=C/C(/C)=C/C=C/C(/C)=C/C=C/C=C(C)C=CC=C(C)C=CC1=C(C)C(=O)[C@@H](O)CC1(C)C MQZIGYBFDRPAKN-ZWAPEEGVSA-N 0.000 title claims abstract description 114
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Abstract
本发明属于调味品的制备技术领域,具体涉及一种含虾青素的调味品及其制备方法。该方法首先采用低速珠磨法破碎酵母细胞壁,使虾青素直接乳化于水中,虾青素浓度可达到1043.17mg/L,避免了乳化剂和有机溶剂的使用。然后添加少量安琪酵母复配酶进行酶解,氨基氮得率高达3.51~3.65%,固形物得率高达47.18~49.22%。最后依次加入糊化的多孔淀粉溶液和明胶,所得含虾青素的微囊粉的包埋率和载药量分别为75.62~88.5%和1.55~10.42mg/g。该微囊粉颜色鲜艳,虾青素稳定性和水溶性较高。其制备方法耗时短,条件温和,可降低红法夫酵母工业运用成本。
Description
技术领域
本发明属于调味品的制备技术领域,具体涉及一种含虾青素的调味品及其制备方法。
背景技术
酵母抽提物含有肽类化合物、人体所需的18种氨基酸、多种核苷酸、糖分及各种微量元素等,是一种兼具调味、营养和保健三大功能的天然复合添加剂。其是以酵母为原料,利用现代生物技术将酵母菌体内的蛋白质和核酸物质降解制备而得。采用的酵母原料中,红发夫酵母的总氮含量4.8%,总碳40%,RNA平均量为8.2%,是一种优选的酵母抽提物原料。由于与其他酵母相比,红发夫酵母胞内富含虾青素,是天然虾青素的来源之一,因此由其制备的酵母抽提物还兼具虾青素的抗氧化及增强免疫力的功效。
为获得红发夫酵母的胞内产物虾青素,在工业化开发过程中必须建立简易且效高的细胞破壁方法。玻璃珠磨是一种操作简单、有效且适合大规模应用的方法。然而现有技术中往往采用高速的珠磨条件来提高破壁效率,其会产生局部高温,导致虾青素损失。此外,由于虾青素不溶于水和大部分有机溶剂,在提取纯化方面目前多使用丙酮和乙醇,会造成丙酮残留和大量的乙醇消耗。
此外,由于虾青素具有长的共轭双键和分子末端羟基酮,其高度不饱和结构导致了其不稳定性,因此对其进行微囊化处理,以期望能达到提高稳定性以及使用过程中缓释的效果。然而,现有研究中往往采用单一壁材,这难以显著提高虾青素稳定性。
发明内容
为克服现有技术的缺点和不足,本发明的首要目的在于提供一种含虾青素的调味品的制备方法。
本发明的另一目的在于提供由上述方法制备而得的含虾青素的调味品。
为实现上述目的,本发明采用的技术方案如下:
一种含虾青素的调味品的制备方法,包括以下步骤:
(1)红发夫酵母细胞的破壁处理
以水为溶剂,配制浓度为10~100g/L的酵母菌悬液,加入粒径为0.8~2.5mm的玻璃珠,使酵母菌悬液中玻璃珠含量为100~500g/L,于120~170rpm条件下震荡破壁,然后移除玻璃珠和沉淀,得到破壁的酵母菌悬液;
(2)酵母抽提物的制备
向破壁的酵母菌悬液中添加安琪酵母复配酶,调节pH值为5~8,进行酶解反应,然后灭酶活,再固液分离,得到含虾青素的酵母抽提物的上清液;
(3)酵母抽提物的微囊化
将多孔淀粉加入水中,加热进行糊化,得到糊化的淀粉溶液,降温至50~60℃,然后将含虾青素的酵母抽提物的上清液加入至糊化的淀粉溶液中,搅拌均匀,再加入固体明胶,震荡,最后真空冷冻干燥,粉碎,过筛,即得含虾青素的调味品;
所述含虾青素的调味品为微囊粉,其芯材为含虾青素的酵母抽提物,其壁材为多孔淀粉和明胶。
步骤(1)中所述的玻璃珠的粒径优选为0.8~1.2mm;更优选为1mm。
步骤(1)中所述的酵母菌悬液中玻璃珠含量优选为300~500g/L;更优选为300g/L。
步骤(1)中所述的震荡破壁的温度优选为22~50℃,更优选为22~40℃。
步骤(1)中所述的震荡破壁的转速优选为150rpm。
步骤(1)中所述的震荡破壁的时长优选为6~48h;更优选为48h。
步骤(2)中所述的安琪酵母复配酶的添加量按其所述的破壁的酵母菌悬液中酵母菌干重的0.5~3wt.%计算。
步骤(2)中所述的酶解反应的温度优选为20~50℃,更优选为40℃。
步骤(2)中所述的酶解反应的时长优选为6~24h。
步骤(2)中所述的固液分离的方式优选为离心。
所述的离心的条件优选为8000~12000rpm离心10~20min。
步骤(3)中所述的水优选为蒸馏水。
步骤(3)中所述的加热的方法优选为沸水浴。
步骤(3)中所述的加热的时长优选为10~20min。
步骤(3)中所述的水的用量优选为按多孔淀粉:水=质量比25:75配比计算。
步骤(3)中所述的芯材与所述的壁材优选为按质量比1:1~1:10.6配比;更优选为按质量比1:1~1:6配比。
步骤(3)中所述的壁材中,多孔淀粉和明胶优选为按质量比1:2~1:5配比,更优选为按2:5配比。
步骤(3)中所述的震荡的温度优选为20~50℃,更优选为40℃。
步骤(3)中所述的震荡的时长优选为2~4h。
步骤(3)中所述的真空冷冻干燥的时长优选为24h。
本发明进一步提供一种由上述方法制备而得而得的含虾青素的调味品。
本发明与现有技术相比,具有如下优点和有益效果:
(1)本发明采用一定粒径、较为温和的珠磨法对酵母进行破壁,由于破壁时间的延长,可直接将虾青素在玻璃珠剪切力的作用下乳化分散于水中,虾青素浓度可达到1043.17mg/L,避免了有机溶剂和乳化剂的使用,从而减少了工艺操作。同时可避免高转速珠磨带来的局部升温导致虾青素损失的问题。
(2)本发明的微囊化处理过程中,首先对多孔淀粉进行了预糊化,并降低多孔淀粉的添加比例,可提高微囊粉的水溶性。首先采用多孔淀粉吸附虾青素,然后添加明胶,以保留虾青素的颜色,遮盖酵母的气味,由多孔淀粉和明胶组成的复合壁材可显著提高虾青素的稳定性和水溶性,有利于酵母抽提物在食品中的应用。由此制备的含虾青素的微囊粉的包埋率为75.62~88.5%,载药量为1.55~10.42mg/g。
(3)本发明制备方法为无添加且无残留的绿色工艺,工艺流程耗时短,条件温和,适于工业化。
具体实施方式
下面结合实施例对本发明作进一步地详细说明,但本发明的实施方式不限于此。对于未特别注明的工艺参数,可参照常规技术进行。
实施例1
本实施例提供采用玻璃珠对红发夫酵母进行破壁的条件探索
(一)不同浓度红发夫酵母的细胞破壁:
向20ml水中加入红发夫酵母(74220,购自ATCC,下同),得到浓度为10、20和40g/L(干重)的酵母菌悬液,向菌悬液中添加粒径为1mm的玻璃珠,使酵母菌悬液中玻璃珠含量为200g/L,然后在摇床中,于22℃和150rpm的搅拌速度下震荡破壁6h。破壁结束后,在10000rpm下离心10min,收集沉淀和上清液。用丙酮提取并以石油醚反萃取上清液中的虾青素(提取方法参照Gil-Hwan An.Applied and environmental microbiology,1988,55(1):116-124),用丙酮提取沉淀中的虾青素,然后采用分光光度计分别检测上清液与沉淀中的虾青素含量。以酸-热处理法提取的虾青素量(提取方法参照Ling-Yan Zhou,International Journal of Modern Biology and Medicine,2015,6(2):136-145)为对照,即默认热酸法提取的虾青素为全部虾青素量,提取率为本方法提取的虾青素总量占全部虾青素量的百分比。
结果如下:酵母浓度为10g/L的菌悬液的虾青素提取率为58.00%;酵母浓度为20g/L的菌悬液的虾青素提取率为64.16%;酵母浓度为40g/L的菌悬液的虾青素提取率为33.13%。
(二)进一步提高红发夫酵母浓度对虾青素溶解度的影响:
向20ml水中加入红发夫酵母,得到浓度为20、50和100g/L(干重)的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为500g/L,然后在摇床中,于22℃和150rpm的搅拌速度下震荡破壁48h。破壁结束后,在10000rpm下离心10min,收集沉淀和上清液。用丙酮提取并以石油醚反萃取上清液中的虾青素,用丙酮提取沉淀中的虾青素,然后采用分光光度计分别检测上清液与沉淀中的虾青素含量。以酸-热处理法提取的虾青素量为对照,即默认热酸法提取的虾青素为全部虾青素量,计算虾青素的提取率。
结果如下:红发夫酵母浓度为20和50g/L的菌悬液的虾青素提取率都可达到100%,破壁后上清液中的虾青素浓度分别为459.57和843.06mg/L;红发夫酵母浓度为100g/L的菌悬液的虾青素提取率为92.36%,破壁后上清液中的虾青素浓度为1043.17mg/L。
(三)不同温度下红发夫酵母的细胞破壁:
向20ml水中加入红发夫酵母,得到浓度为20g/L(干重)的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为200g/L,然后在摇床中,在150rpm的搅拌速度下,于22℃、40℃和50℃下震荡破壁6h。破壁结束后,在10000rpm下离心10min,收集沉淀和上清液。用丙酮提取并以石油醚反萃取上清液中的虾青素,用丙酮提取沉淀中的虾青素,然后采用分光光度计分别检测上清液与沉淀中的虾青素含量。以酸-热处理法提取的虾青素量为对照,即默认热酸法提取的虾青素为全部虾青素量,计算虾青素的提取率。
结果如下:22℃虾青素提取率为64.16%;40℃虾青素提取率为67.83%;50℃虾青素提取率为36.01%。
(四)不同玻璃珠添加量下红发夫酵母的细胞破壁:
向20ml水中加入红发夫酵母,得到浓度为20g/L(干重)的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量分别为100g/L、200g/L和300g/L,然后在摇床中,于22℃和150rpm的搅拌速度下震荡破壁6h。破壁结束后,在10000rpm下离心10min,收集沉淀和上清液。用丙酮提取并以石油醚反萃取上清液中的虾青素,用丙酮提取沉淀中的虾青素,然后采用分光光度计分别检测上清液与沉淀中的虾青素含量。以酸-热处理法提取的虾青素量为对照,即默认热酸法提取的虾青素为全部虾青素量,计算虾青素的提取率。
结果如下:玻璃珠添加量为100g/L的虾青素提取率为47.66%;玻璃珠添加量为200g/L的虾青素提取率为64.16%;玻璃珠添加量为300g/L的虾青素提取率为96.68%。
(五)不同破壁时间下红发夫酵母的细胞破壁:
向20ml水中加入红发夫酵母,得到浓度为20g/L(干重)的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为500g/L,然后在摇床中,于22℃和150rpm的搅拌速度下分别震荡破壁6h、24h和48h。破壁结束后,在10000rpm下离心10min,收集沉淀和上清液。用丙酮提取并以石油醚反萃取上清液中的虾青素,用丙酮提取沉淀中的虾青素,然后采用分光光度计分别检测上清液与沉淀中的虾青素含量。以酸-热处理法提取的虾青素量为对照,即默认热酸法提取的虾青素为全部虾青素量,计算虾青素的提取率。
结果如下:破壁时间为6、24和48h的虾青素提取率都可达到100%,破壁后上清中的虾青素浓度分别为93.72、306.43和459.57mg/L。
(六)红发夫酵母的珠磨破壁
取1ml浓度为20g/L的酵母悬液于配套的高强度的2mL离心管中,并加入750mg直径为0.1mm的玻璃珠。摇晃混匀,然后放置于Mini-Beadbeater-16珠磨式研磨器(115-230V,50-60Hz)中,在150rpm条件下进行破碎,每次破碎30s,破碎次数分别为8、12和16。
结果如下:破碎8次虾青素提取率为11.84%;破碎12次虾青素提取率为45.07%;破碎16次虾青素提取率为72.13%,但此时沉淀中已洗至无色,说明部分色素损失。
(七)红发夫酵母的酶解破壁
向20ml水中加入红发夫酵母,得到浓度为20g/L(干重)的酵母菌悬液,其中添加安琪酵母复配酶的量分别为0.5wt.%、1wt.%和3wt.%(干重),放置于50℃、150rpm,避光条件下酶解4h。反应结束后,计算虾青素提取率。
结果如下:添加安琪酵母复配酶的量为0.5wt.%,虾青素提取率为23.41%;添加安琪酵母复配酶的量为1wt.%,虾青素提取率为31.70%;添加安琪酵母复配酶的量为3wt.%,虾青素提取率为42.83%。
实施例2
安琪酵母复配酶和玻璃珠同时具有破碎酵母的功能,因此本实施例对比了破壁后酶解与同时破壁酶解的酵母抽提物制备方法:
实验组一:同时破壁酶解
向20ml水中加入红发夫酵母,得到浓度(干重)为20g/L的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为300g/L,再分别添加安琪酵母复配酶0.5wt.%、1wt.%和3wt.%,然后调节菌悬液的pH为8,在40℃,150rpm条件下震荡30h,最后在10000rpm下离心10min,收集上清液A。
实验组二:破壁后酶解
向20ml水中加入红发夫酵母,得到浓度(干重)为20g/L的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为300g/L,然后在摇床中,于40℃和150rpm的条件下震荡破壁6h。过滤出玻璃珠和沉淀,得到破壁的酵母菌悬液。调节酵母菌悬液pH为8,分别添加安琪酵母复配酶0.5wt.%、1wt.%和3wt.%,然后继续在40℃,150rpm条件下震荡酶解24h,最后在10000rpm下离心10min,收集上清液B。
采用甲醛滴定法测定上清液A和上清液B的氨基氮得率(测定方法参照Ri-WangLi.Materials Science and Engineering C,2017,77:1035–1043),取一定量的上清液于105℃下烘至恒重,计算得到固形物含量(固形物含量为固形物占原酵母总质量的百分比)。
结果如下:实验组一中安琪酵母复配酶添加量0.5wt.%、1wt.%和3wt.%(酵母干重)对应得到氨基酸得率分别为2.43、2.61和2.76%,固形物含量分别为39.38、36.25和40.30%。实验组二中安琪酵母复配酶添加量为0.5wt.%、1wt.%和3wt.%(干重)对应得到氨基酸得率分别为3.51、3.58和3.65%,固形物含量分别为48.63、49.22和47.18%,均于高于实验组一的氨基酸得率和固形物含量。因此,采用破壁后酶解的技术方案可获得更好的破壁效果。此外,由于不同复配酶添加量下的氨基酸得率和固形物含量相差不大,因此确定破壁后移除玻璃珠,再添加0.5%安琪酵母复配酶制备酵母抽提物。
实施例3
本实施例提供一种含虾青素的调味品的制备方法及微胶囊制备的对照例:
(1)向20ml水中加入红发夫酵母,得到浓度(干重)为20g/L的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为300g/L,然后在摇床中,于40℃和150rpm的条件下震荡破壁6h。过滤出玻璃珠和沉淀,得到破壁的酵母菌悬液。调节酵母菌悬液pH为8,添加酵母复配酶0.5wt.%,然后继续在40℃,150rpm条件下震荡酶解24h,再对酵母菌悬液煮沸3min灭酶活,在10000rpm下离心10min,得到含虾青素的酵母抽提物的上清液,其虾青素浓度为93.72mg/L。
(2)将多孔淀粉加入到蒸馏水中,浓度为25wt.%,沸水浴10min进行糊化,得到糊化的淀粉溶液,接着,降温至50~60℃。将步骤(1)制备的含虾青素的酵母抽提物的上清液加入到糊化的淀粉溶液中,搅拌吸附20min,然后加入明胶,于不同温度条件下振荡一定时间进行微囊化,形成一种稳定的乳浊液。
其中,采用正交实验设计探究影响色素微囊化的因素,正交试验设计因素和水平如下:上清液:20、30和40ml;明胶添加量:0.7、1.0和1.5g;多孔淀粉添加量:0.5、0.6和0.7g;封装温度:40、45和50℃;封装时间:2、3和4h。
本实施例筛选出最佳比例,上清液(V,ml):明胶(W,g):多孔淀粉(W,g)=400:15:6(芯材(W,g):壁材(W,g)=3.95:21),微囊化温度40℃,微囊化时间2h。包埋率是通过检测微囊粉中色素的总含量和表面的色素量计算出来(参照Gomez-Estaca J.FoodHydrocolloids,2016,61:155-162)。表面色素量通过直接加入丙酮提取得出;微囊粉中色素的总含量,则使微囊粉溶于蒸馏水中,振荡后加入丙酮,混合均匀后离心,采用石油醚反萃取色素,取上层色素-石油醚于分光光度计测吸光度A值;如果下层丙酮中色素提取不完全,再重复加入石油醚,直至下层有机溶剂无色。载药量即每克微囊粉所含色素量。溶解度采用重量法测定(测定方法参照Talita A.Comunian.Food Research International,2013,52(1):373-379)
所得含虾青素的微囊粉的包埋率,载药量和溶解度分别为88.5%,1.55mg/g和79.13%。微囊粉中氨基氮含量测定为1.35%。
(3)将多孔淀粉和明胶同时加入到步骤(1)制备的含虾青素的酵母抽提物的上清液中,搅拌吸附20min,然后于40℃下振荡进行微囊化2h。其中,上清液(V,ml):明胶(W,g):多孔淀粉(W,g)=400:15:6。
结果如下:微囊化后多孔淀粉沉淀在底部,不能形成稳定的乳浊液;得到的微囊粉颜色深浅分布不均匀,包埋率差异较大。对比步骤(2)和步骤(3)的结果可知,采用先加入糊化的多孔淀粉,后加入明胶的微囊化添加顺序更有利于虾青素的微囊化。
实施例4
本实施例提供不同浓度虾青素的微胶囊的制备方法:
向20ml的水中加入红发夫酵母,得到浓度为20、50和100g/L(干重)的酵母菌悬液,向菌悬液中添加1mm玻璃珠,使酵母菌悬液中玻璃珠含量为500g/L,然后在摇床中,于22℃和150rpm的搅拌速度下震荡破壁48h。过滤出玻璃珠和沉淀,得到破壁的酵母菌悬液。调节酵母菌悬液pH为8,添加酵母复配酶0.5%,然后继续在40℃,150rpm条件下震荡酶解24h,再对酵母菌悬液煮沸3min灭酶活,在10000rpm下离心10min,得到含虾青素的酵母抽提物的上清液。上清液按照正交实验微囊化条件,上清液(V,ml):明胶(W,g):多孔淀粉(W,g)=400:15:6,微囊化温度40℃,微囊化时间2h,进行包埋。
结果如下:酵母浓度为20g/L,虾青素包埋率和载药量为87.8%和3.45mg/g;酵母浓度为50g/L,虾青素包埋率和载药量为78.69%和6.79mg/g;酵母浓度为100g/L,虾青素包埋率和载药量为75.62%和10.42mg/g。包埋率和载药量的检测方法参照实施例3。
上述实施例为本发明较佳的实施方式,但本发明的实施方式并不受上述实施例的限制,其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化,均应为等效的置换方式,都包含在本发明的保护范围之内。
Claims (10)
1.一种含虾青素的调味品的制备方法,其特征在于,包括以下步骤:
(1)红发夫酵母细胞的破壁处理
以水为溶剂,配制浓度为10~100g/L的酵母菌悬液,加入粒径为0.8~2.5mm的玻璃珠,使酵母菌悬液中玻璃珠含量为100~500g/L,于120~170rpm条件下震荡破壁,然后移除玻璃珠和沉淀,得到破壁的酵母菌悬液;
(2)酵母抽提物的制备
向破壁的酵母菌悬液中添加安琪酵母复配酶,调节pH值为5~8,进行酶解反应,然后灭酶活,再固液分离,得到含虾青素的酵母抽提物的上清液;
(3)酵母抽提物的微囊化
将多孔淀粉加入水中,加热进行糊化,得到糊化的淀粉溶液,降温至50~60℃,然后将含虾青素的酵母抽提物的上清液加入至糊化的淀粉溶液中,搅拌均匀,再加入固体明胶,震荡,最后真空冷冻干燥,粉碎,过筛,即得含虾青素的调味品;
所述的含虾青素的调味品为微囊粉,其芯材为含虾青素的酵母抽提物,其壁材为多孔淀粉和明胶。
2.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:
步骤(1)中所述的玻璃珠的粒径为0.8~1.2mm;
步骤(1)中所述的酵母菌悬液中玻璃珠含量为300~500g/L。
3.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于,步骤(1)中所述的震荡破壁的条件为:温度是22~50℃,转速为150rpm,破壁时长为6~48h。
4.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:步骤(2)中所述的安琪酵母复配酶的添加量按其为所述的破壁的酵母菌悬液中酵母菌干重的0.5~3wt.%计算。
5.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:步骤(2)中所述的酶解反应的条件为于20~50℃酶解6~24h。
6.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:
步骤(2)中所述的固液分离的方式为离心;
步骤(3)中所述的水为蒸馏水;
步骤(3)中所述的加热的方法为沸水浴。
7.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:
步骤(3)中所述的水的用量按多孔淀粉:水=质量比25:75配比计算。
8.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:
步骤(3)中所述的芯材与所述的壁材按质量比1:1~1:10.6配比;
步骤(3)中所述的壁材中,多孔淀粉和明胶按质量比1:2~1:5配比。
9.根据权利要求1所述含虾青素的调味品的制备方法,其特征在于:
步骤(3)中所述的震荡的温度为20~50℃;
步骤(3)中所述的震荡的时长为2~4h;
步骤(3)中所述的真空冷冻干燥的时长为24h。
10.一种含虾青素的调味品,其特征在于:由权利要求1~9任一项所述的制备方法得到。
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