CN115216462A - 一种用于制备黑水虻油的复合酶制剂及其应用 - Google Patents
一种用于制备黑水虻油的复合酶制剂及其应用 Download PDFInfo
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Abstract
本发明属于昆虫蛋白利用技术领域,具体提供了一种用于制备黑水虻油的复合酶制剂及其应用。本发明提供的这种用于制备黑水虻油的复合酶制剂,包括:木瓜蛋白酶、碱性蛋白酶、酸性蛋白酶、碱性脂肪酶、壳聚糖酶;所述木瓜蛋白酶、所述碱性蛋白酶、所述酸性蛋白酶、所述碱性脂肪酶和所述壳聚糖酶的质量比为(0.5‑3):(3‑6):(0.5‑1.5):(0.1‑1):0.5。利用酶解转化的方式制备黑水虻油,具有较大的经济效益和社会效益。采用本发明提供的复合酶制剂和方法制备的黑水虻油中脂肪酸含量丰富,中链脂肪酸含量高,具有抗菌效果,能够减少养殖过程中抗生素的使用,此外,酶解工艺处理后黑水虻油中月桂酸含量提高,为开发昆虫油脂资源在水产饲料应用方面奠定基础。
Description
技术领域
本发明属于昆虫蛋白利用技术领域,具体涉及一种用于制备黑水虻油的复合酶制剂及其应用。
背景技术
随着畜禽、水产养殖业的发展,饲料资源相对短缺的状况日益严重,脂肪作为三大营养物质之一,在水产动物生长发育过程中十分重要。当前饲料油脂添加量不断增加,鱼油作为水产饲料行业的主要油脂源目前已出现供不应求的现象,因此开发寻找适宜的油脂资源也越来越被重视。昆虫中油脂含量丰富,脂肪酸组成合理,大多包含软脂酸以下不饱和脂肪酸,消化性能较好,且油酸和亚油酸含量丰富,有的昆虫亚麻酸含量也很高,亚油酸及亚麻酸为淡水鱼类的必需脂肪酸,因此,昆虫油脂是一种潜在的水产饲料用脂肪来源。但目前针对昆虫油的研究不多,相关研究还不够深入,因此限制了昆虫油脂在饲料行业的有效应用。
黑水虻虫粉中干物质含量≥92%,粗蛋白含量约45%,粗脂肪含量约33%,钙含量约0.96%,总磷含量约0.80%。此外,黑水虻粗脂肪中,中链脂肪酸与多不饱和脂肪酸含量占脂肪酸总量的60%以上,其中月桂酸含量占脂肪酸的40%以上。尽管黑水虻富含丰富的营养物质,但关于黑水虻产品的研究相对较少。黑水虻虫蛹含17.63%的亚油酸和32.03%的不饱和脂肪酸,多不饱和脂肪酸与单不饱和脂肪比例约为1.36,可作为优质的脂肪酸源,且提取的黑水虻油具有更强的抗氧化能力。许多国家都把人工饲养昆虫蛋作为解决蛋白饲料来源的主攻方向,黑水虻等资源昆虫因其生长速度快、高蛋白、高脂肪且含抗菌肽等特点而受到广泛关注。
发明内容
本发明的目的是资源化利用将黑水虻,扩充饲料原料来源。
为此,本发明提供了一种用于制备黑水虻油的复合酶制剂,包括:木瓜蛋白酶、碱性蛋白酶、酸性蛋白酶、碱性脂肪酶、壳聚糖酶;所述木瓜蛋白酶、所述碱性蛋白酶、所述酸性蛋白酶、所述碱性脂肪酶和所述壳聚糖酶的质量比为(0.5-3):(3-6):(0.5-1.5):(0.1-1):0.5。
具体的,上述木瓜蛋白酶的酶活力不低于8000U/g,碱性蛋白酶的酶活不低于200000U/g,酸性蛋白酶的酶活力不低于100000U/g,碱性脂肪酶的酶活力不低于80000U/g,壳聚糖酶的酶活力不低于8000U/g。
本发明还提供了一种黑水虻油的制备方法,包括以下步骤:
(1)按比例配制上述复合酶制剂备用;
(2)选取黑水虻幼虫,超声诱导后饲养;
(3)将饲养后的黑水虻去水除杂后灭活;
(4)向灭活后的黑水虻中加入柠檬酸调节pH至7.0-7.5,随后打成黑水虻虫浆;
(5)将复合酶制剂加入黑水虻虫浆中进行酶解,得到黑水虻酶解液;
(6)采用超临界CO2萃取法对黑水虻酶解液进行脱油处理,提取上清烘干得到黑水虻油成品。
具体的,上述步骤(2)中超声诱导条件为:用功率为100W超声波进行超声诱导20min。
具体的,上述步骤(3)中灭活方法为:90-95℃热水处理5-20min;水与黑水虻的质量比为(0.5-1):1。
具体的,上述步骤(4)中柠檬酸为质量分数为10%的柠檬酸水溶液,黑水虻浓度为(10-20):100w/v。
具体的,上述步骤(5)中复合酶制剂的添加量为黑水虻质量的4.5-10%。
具体的,上述步骤(5)中酶解条件为:40-60℃,80~120rpm/min下酶解8-16h。
具体的,上述步骤(6)的超临界CO2萃取法中温度为40-60℃,压力15-20Mpa,干燥温度为60℃。
与现有技术相比,本发明具有以下优点和有益效果:
本发明提供的这种用于制备黑水虻油的复合酶制剂通过不同蛋白酶与壳聚糖酶进行复配后,能够有效降解大分子蛋白及几丁质、壳聚糖,蛋白酶能随机水解大分子的蛋白质中的肽键,配合外切和内切蛋白酶生成α-氨基酸、二肽或三肽等短链蛋白质小肽类物质提供氮源,同时也能将几丁质与蛋白和脂肪交联的物质进行有效水解,通过酶制剂各单酶之间的协同效果,有效降解几丁质、蛋白及脂肪偶联物,效果明显优于单酶效果,水解效率高;通过对酶的配比与调控,酶解作用不仅仅使得有效成分总量大幅度上升,而且产物得率更高效果更稳定。本发明利用酶解转化的方式制备黑水虻油,为昆虫油脂资源的开发利用及缓解饲料行业油源供应不足的情况提供参考资料,具有较大的经济效益和社会效益。采用本发明提供的复合酶制剂和方法制备的黑水虻油中脂肪酸含量丰富,C4~C18的比例高达99%,其中C4~C10的比例为5~15%、C12的比例为30-50%、C14~C18的比例为45~55%,中链脂肪酸含量高,具有抗菌效果,能够减少养殖过程中抗生素的使用。此外,复合酶配伍不同,实际产物不同,采用气相色谱法测得酶解工艺处理后黑水虻油中月桂酸含量(C12:0)提高至36-38%左右,为开发昆虫油脂资源在水产饲料应用方面奠定基础,另外产物抗菌能力强,除脂肪酸具有抑菌功能外,富含小分子活性肽具有较强抗菌能力,酶解充分、产物丰富,复合抗菌产品,应用前景较大。
以下将结合附图对本发明做进一步详细说明。
附图说明
图1是本发明实施例4中大肠杆菌抑菌图;其中a为实施例2抑菌结果;b为实施例3抑菌结果;c为阳性对照结果;d为阴性对照结果;e为比较例结果。
图2是本发明实施例4中沙门氏菌抑菌图;其中a为实施例2抑菌结果;b为实施例3抑菌结果;c为阳性对照结果;d为阴性对照结果;e为比较例结果。
具体实施方式
下面将结合实施例对本发明中的技术方案进行清楚、完整地描述,显然,所描述的实施例仅仅是本发明一部分实施例,而不是全部的实施例。尽管已经详细描述了本发明的代表性实施例,但是本发明所属技术领域的普通技术人员将理解,在不脱离本发明范围的情况下可以对本发明进行各种修改和改变。因此,本发明的范围不应局限于实施方案,而应由所附权利要求及其等同物来限定。
下面通过具体实施例对本发明的用于制备黑水虻油的复合酶制剂及其应用的效果进行研究。
实施例1:
本实施例提供了一种用于制备黑水虻油的复合酶制剂,包括:酶活为8000U/g的木瓜蛋白酶、酶活为200000U/g的碱性蛋白酶、酶活为100000U/g的酸性蛋白酶、酶活为80000U/g的碱性脂肪酶、酶活为8000U/g的壳聚糖酶;所述木瓜蛋白酶、所述碱性蛋白酶、所述酸性蛋白酶、所述碱性脂肪酶和所述壳聚糖酶的质量比为0.5:5:1:0.1。
实施例2:
本实施例采用实施例1提供的复合酶制剂制备黑水虻油,具体步骤如下:
(1)选取黑水虻幼虫5龄幼虫,用功率为100W超声波进行超声诱导20min,诱导后继续常规饲养48h;
(2)去水清除黑水虻幼虫中杂质,90℃热水处理5min;水与黑水虻的质量比为0.5:1;
(3)向灭活后的黑水虻中加入质量分数为10%的柠檬酸水溶液调节pH至7.0,随后打成黑水虻虫浆,黑水虻幼虫底物浓度为10:100w/v;
(4)以黑水虻质量计,将4.5%的复合酶制剂加入黑水虻虫浆中进行酶解,酶解温度45℃,转速80rpm/min,酶解时间8h。,得到黑水虻酶解液;
(5)采用超临界CO2萃取法对黑水虻酶解液进行脱油处理,提取上清烘干得到黑水虻油成品,超临界CO2萃取法中温度为40℃,压力15Mpa,干燥温度为60℃。
检测黑水虻油成品中脂肪酸组成,结果如表1所示。
实施例3:
本实施例采用实施例1提供的复合酶制剂制备黑水虻油,具体步骤如下:
(1)选取黑水虻幼虫5龄幼虫,用功率为100W超声波进行超声诱导20min,诱导后继续常规饲养48h;
(2)去水清除黑水虻幼虫中杂质,95℃热水处理20min;水与黑水虻的质量比为1.0:1;
(3)向灭活后的黑水虻中加入质量分数为10%的柠檬酸水溶液调节pH至7.5,随后打成黑水虻虫浆,黑水虻幼虫底物浓度为20:100w/v;
(4)以黑水虻质量计,将10%的复合酶制剂加入黑水虻虫浆中进行酶解,酶解温度60℃,转速120rpm/min,酶解时间16h,得到黑水虻酶解液;
(5)采用超临界CO2萃取法对黑水虻酶解液进行脱油处理,提取上清烘干得到黑水虻油成品,超临界CO2萃取法中温度为60℃,压力20Mpa,干燥温度为60℃。
检测本实施例制备的黑水虻油成品和市售黑水虻油中脂肪酸组成,结果如表1所示。
表1黑水虻油脂肪酸组成
实施例4
采用牛津杯法测定实施例2和实施例3制备的黑水虻油对大肠杆菌致病菌和沙门氏菌致病菌的抑菌效果,本实施例所用致病菌来自中国工业微生物菌种保藏中心,大肠杆菌致病菌编号为CICC10305,沙门氏菌致病菌编号为CICC21493。每个抑菌实验分别重复3次,其中大肠杆菌阳性对照采用氨苄青霉素,沙门氏菌阳性对照采用环丙沙星,阴性对照均为水。抑菌结果如图1-2所示,从抑菌平板上可以看到,实施例2(a)和实施例3(b)的抑菌效果明显比为比较例市售商品黑水虻油(e)抑菌效果好。
以上例举仅仅是对本发明的举例说明,并不构成对本发明的保护范围的限制,凡是与本发明相同或相似的设计均属于本发明的保护范围之内。
Claims (10)
1.一种用于制备黑水虻油的复合酶制剂,其特征在于,包括:木瓜蛋白酶、碱性蛋白酶、酸性蛋白酶、碱性脂肪酶、壳聚糖酶;所述木瓜蛋白酶、所述碱性蛋白酶、所述酸性蛋白酶、所述碱性脂肪酶和所述壳聚糖酶的质量比为(0.5-3):(3-6):(0.5-1.5):(0.1-1):0.5。
2.如权利要求1所述的用于制备黑水虻油的复合酶制剂,其特征在于:所述木瓜蛋白酶的酶活力不低于8000U/g,所述碱性蛋白酶的酶活不低于200000U/g,所述酸性蛋白酶的酶活力不低于100000U/g,所述碱性脂肪酶的酶活力不低于80000U/g,所述壳聚糖酶的酶活力不低于8000U/g。
3.一种黑水虻油的制备方法,其特征在于,包括以下步骤:
(1)按比例配制权利要求1-2任意一项所述的复合酶制剂备用;
(2)选取黑水虻幼虫,超声诱导后饲养;
(3)将饲养后的黑水虻去水除杂后灭活;
(4)向灭活后的黑水虻中加入柠檬酸调节pH至7.0-7.5,随后打成黑水虻虫浆;
(5)将复合酶制剂加入黑水虻虫浆中进行酶解,得到黑水虻酶解液;
(6)采用超临界CO2萃取法对黑水虻酶解液进行脱油处理,提取上清烘干得到黑水虻油成品。
4.如权利要求3所述的黑水虻油的制备方法,其特征在于,所述步骤(2)中超声诱导条件为:用功率为100W超声波进行超声诱导20min。
5.如权利要求3所述的黑水虻油的制备方法,其特征在于,所述步骤(3)中灭活方法为:90-95℃热水处理5-20min;水与黑水虻的质量比为(0.5-1):1。
6.如权利要求3所述的黑水虻油的制备方法,其特征在于:所述步骤(4)中柠檬酸为质量分数为10%的柠檬酸水溶液,黑水虻虫浆浓度为(10-20):100w/v。
7.如权利要求3所述的黑水虻油的制备方法,其特征在于:所述步骤(5)中复合酶制剂的添加量为黑水虻质量的4.5-10%;酶解条件为40-60℃,80~120rpm/min下酶解8-16h。
8.如权利要求3所述的黑水虻油的制备方法,其特征在于:所述步骤(6)的超临界CO2萃取法中温度为40-60℃,压力15-20Mpa,干燥温度为60℃。
9.一种黑水虻油,其特征在于:所述黑水虻油通过权利要求3-8任意一项所述制备方法制备。
10.如权利要求9所述黑水虻油在水产饲料中的应用。
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Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN115769852A (zh) * | 2022-12-13 | 2023-03-10 | 珠海市德海生物科技有限公司 | 一种黑水虻幼虫虫粉的加工方法及其应用 |
CN117070274A (zh) * | 2023-09-27 | 2023-11-17 | 深圳市中兴恒熙环保有限公司 | 一种利用黑水虻幼虫制备油脂的方法 |
CN117100674A (zh) * | 2023-10-25 | 2023-11-24 | 广州安芮洁环保科技有限公司 | 一种黑水虻提取物及其在制备具有抗菌作用的产品中的应用 |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20120144732A1 (en) * | 2010-03-08 | 2012-06-14 | Sun Yat-Sen University | Biodiesel and Preparation Method Thereof |
CN105456155A (zh) * | 2015-11-23 | 2016-04-06 | 太仓安佑生物科技有限公司 | 一种以黑水虻幼虫制备化妆品原料的方法及其应用 |
-
2022
- 2022-06-28 CN CN202210747356.6A patent/CN115216462B/zh active Active
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20120144732A1 (en) * | 2010-03-08 | 2012-06-14 | Sun Yat-Sen University | Biodiesel and Preparation Method Thereof |
CN105456155A (zh) * | 2015-11-23 | 2016-04-06 | 太仓安佑生物科技有限公司 | 一种以黑水虻幼虫制备化妆品原料的方法及其应用 |
Non-Patent Citations (3)
Title |
---|
张佰帅等: "动物油脂提取及加工技术研究进展", 油脂加工, vol. 35, no. 12, pages 8 - 11 * |
欧春艳等: "壳聚糖降解研究的最新进展", 广州化工, vol. 41, no. 6, pages 13 - 15 * |
贾申宗等: "黑水虻油替代豆油对淇河鲫生长性能、血清生化指标和肠道消化酶活力的影响", 河南师范大学学报(自然科学版), vol. 50, no. 2, pages 129 - 135 * |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN115769852A (zh) * | 2022-12-13 | 2023-03-10 | 珠海市德海生物科技有限公司 | 一种黑水虻幼虫虫粉的加工方法及其应用 |
CN117070274A (zh) * | 2023-09-27 | 2023-11-17 | 深圳市中兴恒熙环保有限公司 | 一种利用黑水虻幼虫制备油脂的方法 |
CN117100674A (zh) * | 2023-10-25 | 2023-11-24 | 广州安芮洁环保科技有限公司 | 一种黑水虻提取物及其在制备具有抗菌作用的产品中的应用 |
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