CN108850778A - 一种长效稳定型生物质抗氧化剂的制备方法 - Google Patents
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Abstract
本发明涉及一种长效稳定型生物质抗氧化剂的制备方法,属于抗氧化剂制备技术领域。本发明以复合酶作为酶解剂,以虾壳作为原料,经酶解后超声辅助提取,得到虾青素,接着将其和海藻酸钠共混后逐滴滴入氯化钙溶液中,形成凝胶包覆层包裹住虾青素,得到虾青素凝胶小球,接着将柠檬汁和酵母粉混合发酵,再将发酵液和虾青素凝胶小球再次发酵,将发酵滤渣紫外灭菌后晾干,即得长效稳定型生物质抗氧化剂,本发明制得的长效稳定型生物质抗氧化剂在基体上固着时间长,抗氧化时效持久,具有广阔的应用前景。
Description
技术领域
本发明涉及一种长效稳定型生物质抗氧化剂的制备方法,属于抗氧化剂制备技术领域。
背景技术
近年来,由于化学抗氧化添加剂的不安全性和食品安全的考虑,化学抗氧化添加剂的应用受到了较大的限制。因此,高效、低毒的天然食品抗氧化剂的开发成为研究热点领域。
目前,已从各种植物、动物组织中提取得到了抗氧化活性显著的物质,如茶叶中的茶多酚、大豆中的异黄酮、枸杞中的多糖等物质,虾青素最早是从虾蟹的壳中分离提取获得,直到上世纪八十年代人们才开始关注虾青素的生物活性功能。
在虾青素特殊分子结构中,共轭双键链及末端的不饱和酮基和羟基相互作用,加剧分子之间的摩擦力度,可以促进电子的自由移动,自由基负责提供电子与未配对电子相互吸引,能有效地猝灭附近的自由基及有害的活性氧,这些结构使得虾青素的抗氧化活性较强,在抗氧化能力方面远远超过其他的类胡萝卜素。但是虾青素稳定性比较差、在高温、潮湿等条件下极易发生变性反应,并且虾青素与基体间附着性差,容易脱落导致抗氧化时间缩短,这使得虾青素的应用受到了很大的影响。
因此为了延长虾青素的抗氧化作用时间和效果,并增加虾青素的稳定性,发明一种长效稳定型生物质抗氧化剂对生物质抗氧化剂制备技术领域具有积极的意义。
发明内容
本发明主要解决的技术问题,针对目前虾青素稳定性比较差、在高温、潮湿等条件下极易发生变性反应,并且虾青素与基体间附着性差,容易脱落导致抗氧化时间缩短的缺陷,提供了一种长效稳定型生物质抗氧化剂的制备方法。
为了解决上述技术问题,本发明所采用的技术方案是:
一种长效稳定型生物质抗氧化剂的制备方法,其特征在于具体制备步骤为:
(1)将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,冷冻,再将冷冻的虾壳放入组织粉碎机中,粉碎30~40min,得到虾壳粉碎物;
(2)将上述虾壳粉碎物和复合酶混合后装入酶解罐中,酶解,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯混合得到混合液;
(3)将上述混合液装入超声振荡仪中,超声振荡反应,得到反应液,再将反应液移入离心机中,离心处理,分离得到上层液,将上层液放入减压蒸馏装置中,蒸馏得到虾青素;
(4)将海藻酸钠和去离子水混合后移入水浴锅,加热升温至40~50℃,用磁力搅拌机以300~400r/min的转速搅拌10~15min,再降低搅拌转速至200~300r/min,在搅拌状态下再加入上述虾青素,继续搅拌10~20min,得到预制液;
(5)将质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以50~60r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;
(6)将柠檬汁和酵母粉混合后装入发酵罐中,发酵,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,二次发酵,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌8~10h,自然晾干后即得长效稳定型生物质抗氧化剂。
步骤(1)中所述的蛋白酶、脂肪酶、几丁质酶和多酚氧化酶的质量比为3:2:1:1,冷冻的温度为-20~-15℃,冷冻的时间为10~12h。
步骤(2)中所述的虾壳粉碎物和复合酶的质量比为10:1,酶解的温度为25~30℃,酶解的时间为6~8h,酶解产物和乙酸乙酯的质量比为1:5。
步骤(3)中所述的超声振荡反应的频率为30~35kHz,超声振荡反应的时间为35~45min,离心处理的转速为4000~4500r/min,离心处理的时间为10~15min,减压蒸馏的压力为1.5~2.0kPa、温度为60~70℃。
步骤(4)中所述的海藻酸钠和去离子水的质量比为1:10,虾青素的加入量为海藻酸钠质量的50%。
步骤(5)中所述的质量分数为90%的氯化钙溶液的加入量为预制液体积的3~5倍。
步骤(6)中所述的柠檬汁和酵母粉的质量比为10:1,发酵的温度为25~35℃,发酵的时间为7~9天,继续发酵的温度为30~35℃,继续发酵的时间为6~8h。
本发明的有益效果是:
(1)本发明以复合酶作为酶解剂,以虾壳作为原料,经酶解后超声辅助提取,得到虾青素,接着将其和海藻酸钠共混后逐滴滴入氯化钙溶液中,形成凝胶包覆层包裹住虾青素,得到虾青素凝胶小球,接着将柠檬汁和酵母粉混合发酵,再将发酵液和虾青素凝胶小球再次发酵,将发酵滤渣紫外灭菌后晾干,即得长效稳定型生物质抗氧化剂,本发明使用的海藻酸钠含有大量的—COO-,在水溶液中可表现出聚阴离子行为,具有一定的黏附性,在酸性条件下,—COO-转变成—COOH,电离度降低,海藻酸钠的亲水性降低,分子链收缩,pH值增加时,—COOH基团不断地解离,海藻酸钠的亲水性增加,分子链伸展,因此,海藻酸钠具有明显的pH敏感性,因此在需抗氧化的基体表面若发生氧化,其表面的pH值只要有些许变化,海藻酸钠就可以感知,并在极其温和的条件下快速形成凝胶,释放抗氧化粉末,起到抗氧化作用,而海藻酸钠 G单元上的Na+与氯化钙的二价钙离子会发生离子交换反应,G单元堆积形成交联网络结构,从而形成水凝胶,而抗氧化钛粉表面的氯化钙固定层由于粒径更大的二价钙离子被替换成了粒径更小的钠离子,因此固定层晶格中的空穴数量增多,这些空穴的产生使得固定层具有更高的物理和化学活性,吸附活性也得到显著提高,因此固定层在保护抗氧化肽粉末的同时,还具备高效的吸附固着能力,使得本申请制得的抗氧化剂能够更加牢固的固着在需抗氧化的基体表面,延长抗氧化剂的抗氧化时间;
(2)本发明还以富含有机羧酸的柠檬汁为原料,在微生物的作用下将有机物柠檬汁降解,产生大量的游离性活性羧基、羟基等,再将富含活性基团的发酵滤液和虾青素凝胶小球进行短时间的二次发酵,将这些活性基团引入到凝胶小球表面,从而使得本发明抗氧化剂可以和基体表面的活性基团产生氢键吸附或者其他化学键合力,这些化学键合力的产生使得本发明看氧化剂和基体之间的固着力得到提高,再一次延长了抗氧化剂的抗氧化时间,具有广阔的应用前景。
具体实施方式
按质量比为3:2:1:1将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,在-20~-15℃下冷冻10~12h,再将冷冻的虾壳放入组织粉碎机中,粉碎30~40min,得到虾壳粉碎物;将虾壳粉碎物和复合酶按质量比为10:1混合后装入酶解罐中,在温度为25~30℃的条件下保温酶解6~8h,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯按质量比为1:5混合得到混合液;将混合液装入超声振荡仪中,以30~35kHz的频率超声振荡反应35~45min,得到反应液,再将反应液移入离心机中,以4000~4500r/min的转速离心处理10~15min,分离得到上层液,将上层液放入减压蒸馏装置中,在压力为1.5~2.0kPa、温度为60~70℃的条件下蒸馏得到虾青素;按质量比为1:10将海藻酸钠和去离子水混合后移入水浴锅,加热升温至40~50℃,用磁力搅拌机以300~400r/min的转速搅拌10~15min,再降低搅拌转速至200~300r/min,在搅拌状态下再加入海藻酸钠质量50%的上述虾青素,继续搅拌10~20min,得到预制液;将预制液体积3~5倍的质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以50~60r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;按质量比为10:1将柠檬汁和酵母粉混合后装入发酵罐中,在25~35℃下发酵7~9天,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,在30~35℃下二次发酵6~8h,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌8~10h,自然晾干后即得长效稳定型生物质抗氧化剂。
按质量比为3:2:1:1将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,在-20℃下冷冻10h,再将冷冻的虾壳放入组织粉碎机中,粉碎30min,得到虾壳粉碎物;将虾壳粉碎物和复合酶按质量比为10:1混合后装入酶解罐中,在温度为25℃的条件下保温酶解6h,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯按质量比为1:5混合得到混合液;将混合液装入超声振荡仪中,以30kHz的频率超声振荡反应35min,得到反应液,再将反应液移入离心机中,以4000r/min的转速离心处理10min,分离得到上层液,将上层液放入减压蒸馏装置中,在压力为1.5kPa、温度为60℃的条件下蒸馏得到虾青素;按质量比为1:10将海藻酸钠和去离子水混合后移入水浴锅,加热升温至40℃,用磁力搅拌机以300r/min的转速搅拌10min,再降低搅拌转速至200r/min,在搅拌状态下再加入海藻酸钠质量50%的上述虾青素,继续搅拌10min,得到预制液;将预制液体积3倍的质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以50r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;按质量比为10:1将柠檬汁和酵母粉混合后装入发酵罐中,在25℃下发酵7天,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,在30℃下二次发酵6h,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌8h,自然晾干后即得长效稳定型生物质抗氧化剂。
按质量比为3:2:1:1将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,在-18℃下冷冻11h,再将冷冻的虾壳放入组织粉碎机中,粉碎35min,得到虾壳粉碎物;将虾壳粉碎物和复合酶按质量比为10:1混合后装入酶解罐中,在温度为28℃的条件下保温酶解7h,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯按质量比为1:5混合得到混合液;将混合液装入超声振荡仪中,以33kHz的频率超声振荡反应40min,得到反应液,再将反应液移入离心机中,以4300r/min的转速离心处理13min,分离得到上层液,将上层液放入减压蒸馏装置中,在压力为1.8kPa、温度为65℃的条件下蒸馏得到虾青素;按质量比为1:10将海藻酸钠和去离子水混合后移入水浴锅,加热升温至45℃,用磁力搅拌机以350r/min的转速搅拌13min,再降低搅拌转速至250r/min,在搅拌状态下再加入海藻酸钠质量50%的上述虾青素,继续搅拌15min,得到预制液;将预制液体积4倍的质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以55r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;按质量比为10:1将柠檬汁和酵母粉混合后装入发酵罐中,在30℃下发酵8天,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,在33℃下二次发酵7h,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌9h,自然晾干后即得长效稳定型生物质抗氧化剂。
按质量比为3:2:1:1将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,在-15℃下冷冻12h,再将冷冻的虾壳放入组织粉碎机中,粉碎40min,得到虾壳粉碎物;将虾壳粉碎物和复合酶按质量比为10:1混合后装入酶解罐中,在温度为30℃的条件下保温酶解8h,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯按质量比为1:5混合得到混合液;将混合液装入超声振荡仪中,以35kHz的频率超声振荡反应45min,得到反应液,再将反应液移入离心机中,以4500r/min的转速离心处理15min,分离得到上层液,将上层液放入减压蒸馏装置中,在压力为2.0kPa、温度为70℃的条件下蒸馏得到虾青素;按质量比为1:10将海藻酸钠和去离子水混合后移入水浴锅,加热升温至50℃,用磁力搅拌机以400r/min的转速搅拌15min,再降低搅拌转速至300r/min,在搅拌状态下再加入海藻酸钠质量50%的上述虾青素,继续搅拌20min,得到预制液;将预制液体积5倍的质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以60r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;按质量比为10:1将柠檬汁和酵母粉混合后装入发酵罐中,在35℃下发酵9天,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,在35℃下二次发酵8h,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌10h,自然晾干后即得长效稳定型生物质抗氧化剂。
对照例以常州市某公司生产的生物质抗氧化剂作为对照例
对本发明制得的长效稳定型生物质抗氧化剂和对照例中的生物质抗氧化剂进行性能检测,检测结果如表1所示:
检测方法,将本发明制得的生物质抗氧化剂和对照例中的抗氧化剂用于生肉保鲜,直接撒在生肉表面,添加量都为100mg/kg,检测两中抗氧剂在生肉上的固着时间,以及可以延长生肉新鲜的时长。
表1 性能检测结果
检测项目 | 实例1 | 实例2 | 实例3 | 对照例 |
固着时间(h) | 195 | 200 | 220 | 72 |
保鲜延长时间(天) | 6 | 6 | 7 | 2 |
由上表中检测数据可以看出,本发明制得的长效稳定型生物质抗氧化剂在基体上固着时间长,抗氧化时效持久,具有广阔的应用前景。
Claims (7)
1.一种长效稳定型生物质抗氧化剂的制备方法,其特征在于具体制备步骤为:
(1)将蛋白酶、脂肪酶、几丁质酶和多酚氧化酶混合得到复合酶,称取虾壳放入冰箱,冷冻,再将冷冻的虾壳放入组织粉碎机中,粉碎30~40min,得到虾壳粉碎物;
(2)将上述虾壳粉碎物和复合酶混合后装入酶解罐中,酶解,酶解结束后得到酶解产物,将酶解产物和乙酸乙酯混合得到混合液;
(3)将上述混合液装入超声振荡仪中,超声振荡反应,得到反应液,再将反应液移入离心机中,离心处理,分离得到上层液,将上层液放入减压蒸馏装置中,蒸馏得到虾青素;
(4)将海藻酸钠和去离子水混合后移入水浴锅,加热升温至40~50℃,用磁力搅拌机以300~400r/min的转速搅拌10~15min,再降低搅拌转速至200~300r/min,在搅拌状态下再加入上述虾青素,继续搅拌10~20min,得到预制液;
(5)将质量分数为90%的氯化钙溶液装入烧杯中,再将烧杯放在摇床上,以50~60r/min的转速进行震荡,在震荡的过程中,再将上述制得的预制液逐滴滴入烧杯中,凝聚成均匀小球,之后过滤分离得到虾青素凝胶小球;
(6)将柠檬汁和酵母粉混合后装入发酵罐中,发酵,发酵结束后过滤分离得到发酵滤液,再将发酵滤液和上述虾青素凝胶小球按等质量比混合后继续放入发酵罐中,二次发酵,发酵结束后过滤分离得到发酵滤渣,放在紫外灯下照射灭菌8~10h,自然晾干后即得长效稳定型生物质抗氧化剂。
2.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(1)中所述的蛋白酶、脂肪酶、几丁质酶和多酚氧化酶的质量比为3:2:1:1,冷冻的温度为-20~-15℃,冷冻的时间为10~12h。
3.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(2)中所述的虾壳粉碎物和复合酶的质量比为10:1,酶解的温度为25~30℃,酶解的时间为6~8h,酶解产物和乙酸乙酯的质量比为1:5。
4.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(3)中所述的超声振荡反应的频率为30~35kHz,超声振荡反应的时间为35~45min,离心处理的转速为4000~4500r/min,离心处理的时间为10~15min,减压蒸馏的压力为1.5~2.0kPa、温度为60~70℃。
5.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(4)中所述的海藻酸钠和去离子水的质量比为1:10,虾青素的加入量为海藻酸钠质量的50%。
6.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(5)中所述的质量分数为90%的氯化钙溶液的加入量为预制液体积的3~5倍。
7.根据权利要求1所述的一种长效稳定型生物质抗氧化剂的制备方法,其特征在于:步骤(6)中所述的柠檬汁和酵母粉的质量比为10:1,发酵的温度为25~35℃,发酵的时间为7~9天,继续发酵的温度为30~35℃,继续发酵的时间为6~8h。
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