CN113546114A - 一种樱花叶细胞水的提取方法和得到的樱花叶细胞水的应用 - Google Patents
一种樱花叶细胞水的提取方法和得到的樱花叶细胞水的应用 Download PDFInfo
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- CN113546114A CN113546114A CN202010333398.6A CN202010333398A CN113546114A CN 113546114 A CN113546114 A CN 113546114A CN 202010333398 A CN202010333398 A CN 202010333398A CN 113546114 A CN113546114 A CN 113546114A
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Abstract
本发明公开了一种樱花叶细胞水的提取方法,通过低温‑真空(减压)提取技术与酶解技术的有机结合,不需要添加任何溶剂,在较低的温度(30℃‑65℃)下能得到纯天然的樱花叶细胞水。通过本发明方法得到的樱花叶细胞水品质高,富含40多种易挥发活性成分,气味芳香柔和,对人体安全性高,具有一定的抑菌效果;可作为绿色天然的原料应用于食品、保健品、药品及化妆品等领域。
Description
技术领域
本发明涉及农产品处理技术领域,特别是涉及一种樱花叶细胞水的提取方法和得到的樱花叶细胞水的应用。
背景技术
樱花树叶是一种传统中药材,具有止咳、平喘、宣肺、润肠、解酒等功效。现代医学证明,樱花树叶富含维生素E、油酸、亚油酸、亚麻酸、棕榈酸、新植二烯、菠菜烯、叶绿醇、黄酮等活性成分,具有防治心脑血管疾病、抑制恶性肿瘤、抗衰老、美容养颜、改善记忆、提高视力、防治肝炎肝硬化、增进食欲、防止化疗引起的脱发、糖尿病及其并发症、过敏性疾病等功效。同时,樱花树叶资源丰富,成本低,具有很大的开发利用前景。
目前的樱花树叶的利用价值还未被完全开发,对于樱花树叶的利用,主要是在食品领域。中国专利CN200810045768.5通过对樱树叶进行深度加工(发酵、干燥等步骤)制得一种干燥樱树叶制品,将其用于制作饮料、冰激凌等。中国专利CN200410059666.0通过有机溶剂萃取法提取得到樱花树叶的乙醇提取物来扩大樱花树叶的利用范围,但由于提取物中含有有机溶剂而使其应用受限。因此,对于樱花树叶的利用还有很大的开发空间。本发明对于樱花树叶的开发利用是以提取樱花叶细胞水的方向来实现。
现有技术中对于植物细胞水的提取,中国专利申请CN109730948A公开了一种采用超声低温旋蒸法和酶法相结合制备牡丹鲜花细胞水的方法:首先通过压榨法得到榨汁和残渣1,再将残渣1旋蒸得到细胞水1和残渣2,最后将残渣2进行酶解后再旋蒸得到细胞水2。将榨汁、细胞水1和细胞水2混合后得到牡丹花细胞水。该方法具有较高的提取效率,但是具有如下缺陷:(1)采用压榨法,后与真空提取法得到的液体混合,这样会带入多糖、色素和刺激气味,导致防腐和脱色问题;细胞水的应用会受到一定的限制;(2)工艺后期配合其他植物一起蒸馏,解决防腐问题,但是容易改变原有的牡丹花细胞水成分和气味,后期生产也无法控制品质。
发明内容
本发明的目的在于,提供一种樱花叶细胞水的提取方法,能够提取得到富含挥发性活性成分、利用价值高的樱花叶细胞水。
本发明是通过以下技术方案实现的:
一种樱花叶细胞水的提取方法,包括以下步骤:不加入溶剂,将樱花叶在温度30℃~65℃、压力-60kPa~-101kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取1.5~2.5小时,得到初提细胞水和初提樱花叶残渣,在初提细胞水中加入初始樱花叶总重量为基准的0.25%~0.45%的纤维素酶和0~0.1%的果胶酶后再倒回初提樱花残渣中,在温度35℃~50℃、压力-85kPa~-101kPa条件下再次提取5~7小时,收集得到樱花叶细胞水。
优选的,所述初步提取的温度为35℃~50℃,压力为-85kPa~-101kPa。优选的温度与压力,细胞水能更好的渗出以及蒸发出来被冷凝收集,同时在较低温度下提取能够减少挥发性活性成分的损失。
本发明通过在初提细胞水中加入一定量的酶,再次投入容器中对樱花叶残渣进行提取,具有如下有益优点:第一、初提细胞水表面张力低,渗透性好;第二、初提细胞水pH为3~7,无需额外调节pH,有利于提高酶活性;第三、酶解能够加速破壁;第四、低温真空技术。通过四种效应的协同,能够在较低温度(35~50℃)下控制酶解速度,加快细胞液流出速度。再次提取步骤中的前1小时左右会先蒸除掉倒回容器内的初提细胞水,在这期间酶解速度快,通过初提液体的量与再次提取的工艺条件来控制酶解的时长(此时初提液体的多少就至关重要,多了会延长酶解时间,使酶解过度;少了会缩短酶解时间,酶解不充分),避免了传统酶解法需要加入大量水而稀释提取液以及酶解过度带来刺激气味的问题。因此,初步提取时间是关键参数之一,如果时间太短,初提樱花叶细胞水过少,加入酶后细胞水很难浸润樱花叶残渣,导致酶解不能正常进行。如果初步提取时间过长,细胞水流出过多,增长了酶解时间,带来过度酶解的风险;同时也降低了后续提取效率。
关于初提樱花叶细胞水的渗透性,通过实验发现,以溶剂法提取樱花叶残渣时,相比于用纯水作为溶剂,采用樱花叶细胞水作为溶剂能够多提取出20%以上的黄酮和多糖。
优选的,纤维素酶的加入量为0.3%~0.4%,果胶酶加入量为0.01%~0.06%。控制酶的加入量,且酶是大分子蛋白质,能够吸附在樱花叶残渣细胞表面,无法在低温真空条件挥发出来,无需再进行后续处理。
本发明所述的樱花叶采用新鲜的樱花树叶,无霉变且未腐烂。新鲜的樱花叶水分饱满,不枯黄;一般为提取当日采摘;或采摘后用冷藏等保鲜手段保存后还保持其新鲜度的。
可将樱花叶切割成宽度为1mm~10mm后再进行提取,有利于提高提取速率和充分提取。
在提取过程中进行搅拌,搅拌速度为1~150转/分钟。防止局部高温;而且再次提取步骤中通过搅拌可以使加入酶的初提细胞水与樱花叶充分混合,保证酶解的进行。
收集过程中进行冷凝,冷凝温度为-10℃~8℃。
本发明的提取方法不需添加其他溶剂,得到的樱花叶细胞水100%是由樱花叶提取出来的,纯度高,绿色天然。
通过本发明的方法提取得到的樱花叶细胞水,可应用于用于食品、保健品、药品及化妆品领域。
本发明与现有技术相比,具有如下有益效果:
本发明通过低温-真空(减压)技术与酶解技术的有机结合来提取樱花叶细胞水,利用初提樱花叶细胞水的高渗透性,将其作为酶解溶剂,达到充分提取的目的。相比于单一的低温-真空提取技术,细胞水的提取率更高且能提取出更多的挥发性活性成分;同时避免了传统酶解法容易酶解过度使提取液具有杂味的问题。
通过本发明方法提取得到的樱花叶细胞水,无色澄清透明;富含40多种挥发性活性成分;气味芳香柔和,安全性高,具有一定的抑菌性;利用价值高,能够应用于食品、保健品、药品及化妆品等领域。
附图说明
图1为实施例1樱花叶细胞水安全性测试数据柱状图。
具体实施方式
下面结合具体实施例对本发明进行详细说明。以下实施例将有助于本领域的技术人员进一步理解本发明,但不以任何形式限制本发明。应当指出的是,对本领域的普通技术人员来说,在不脱离本发明构思的前提下,还可以做出若干变形和改进。这些都属于本发明的保护范围。
实施例和对比例采用新鲜的樱花树叶,无霉变且未腐烂。洗净沥干水后,将其切割成宽度为4mm~5mm后进行提取实验。
实施例1:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在40℃,压力-90kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取1.5小时,得到初提细胞水和初提残渣,在初提细胞水中加入160g纤维素酶和25g果胶酶后再倒入初提残渣中,在40℃、压力-90kPa下再次提取6小时;全程搅拌为45转/分,冷凝温度为-8℃;收集得到35.3kg樱花叶细胞水,无色澄清透明,气味芳香柔和。提取率为70.6%。
实施例2:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在30℃,压力-100kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取2小时,得到初提细胞水和初提残渣,在初提细胞水中加入125g纤维素酶和30g果胶酶后再倒入初提残渣中,在35℃、压力-100kPa下再次提取7小时;全程搅拌为30转/分,冷凝温度为-8℃;收集得到36.6kg樱花叶细胞水,无色澄清透明,气味芳香柔和。提取率为73.2%。
实施例3:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在65℃,压力-70kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取1.5小时,得到初提细胞水和初提残渣,在初提细胞水中加入175g纤维素酶和20g果胶酶后再倒入初提残渣中,在50℃、压力-85kPa下再次提取5小时;全程搅拌为60转/分,冷凝温度为-8℃;收集得到33.9kg樱花叶细胞水,无色澄清透明,气味芳香柔和。提取率为67.8%。
实施例4:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在55℃,压力-80kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取1.5小时,得到初提细胞水和初提残渣,在初提细胞水中加入160g纤维素酶和25g果胶酶后再倒入初提残渣中,在40℃、压力-90kPa下再次提取6小时;全程搅拌为45转/分,冷凝温度为-8℃;收集得到35.1kg樱花叶细胞水,无色澄清透明,气味芳香柔和。提取率为70.2%。
实施例5:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在40℃,压力-90kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取2.5小时,得到初提细胞水和初提残渣,在初提细胞水中加入160g纤维素酶和25g果胶酶后再倒入初提残渣中,在40℃、压力-90kPa下再次提取6小时;全程搅拌为45转/分,冷凝温度为-8℃;收集得到36.0kg樱花叶细胞水,无色澄清透明,气味芳香柔和。提取率为72.0%。
对比例1:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在40℃、-90kPa下提取,细胞水形成蒸气后冷凝收集液体,提取7.5小时,全程搅拌为45转/分,冷凝温度-8℃,得到29.7kg樱花叶细胞水,液体无色澄清透明,香味淡。提取率为59.4%。
对比例2:
将50kg樱花叶投入150L低温提取设备,与5kg水、160g纤维素酶和25g果胶酶混合后在40℃下搅拌6.5小时,搅拌速度为45转/分,结束后过滤(双重过滤:先离心机过滤,再采用0.22μm滤膜过滤)。得到37.8kg樱花叶细胞水(含有额外添加的5kg水),液体为浅黄色,香味淡且具有明显异味。提取率为65.6%(减去5kg水的重量)。
对比例3:
将50kg樱花叶投入150L低温提取设备,与5kg水、160g纤维素酶和25g果胶酶混合后在40℃下搅拌50分钟,后在40℃、-90kPa下提取7.5小时,樱花叶细胞水形成蒸气后冷凝收集液体,全程搅拌为45转/分,冷凝温度-8℃,收集得到39.1kg樱花叶细胞水(含有额外添加的5kg水),无色澄清透明,香味清淡,有杂味。提取率为68.2%(减去5kg水的重量)。
对比例4:
将50kg樱花叶投入150L低温提取设备,不加入任何溶剂,在40℃,压力-90kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取0.5小时,得到初提细胞水和初提残渣,在初提细胞水中加入160g纤维素酶和25g果胶酶后再倒入初提残渣中,在40℃、压力-90kPa下再次提取6小时;全程搅拌为45转/分,冷凝温度为-8℃;收集得到32.6kg樱花叶细胞水,无色澄清透明,香味较淡。提取率为65.2%。
各项测试方法:
1、樱花叶细胞水挥发性活性成分分析:在80℃进样温度下进行顶空气质检测。(1)仪器信息:Agilent 7980A GC;MS:5975C;50/30μm CAR/PDMS/DVB萃取纤维头,美国SUPELCO公司。
(2)GC-MS条件:色谱柱为HP-INNOWAX毛细管柱子(30m×0.25mm×0.25μm);载气为He,流速1mL/min,分离比5:1;进样温度为250℃;升温程序为起始温度为40℃,保持5min,以8℃/min,升至250℃,保持5min。
质谱条件:EI电离源,能量70eV;离子源温度230℃,四极杆温度150℃,接口温度250℃,扫描范围30-400m/z。
(3)样品前处理:将5mL样品、1g NaCl置于20ml顶空瓶中,拧紧瓶盖。于搅拌模式80℃下平衡5min后,用固相微萃取针80℃下萃取5min,然后于进样口解析5min。
提供实施例1樱花叶细胞水挥发性活性成分分析结果数据,见表1。
表1:实施例1樱花叶细胞水挥发性活性成分分析结果(匹配度和相对含量低的成分不列出)
2、樱花叶细胞水喷雾防腐挑战测试
将实施例和对比例提取得到的樱花叶细胞水,添加到如下喷雾配方中(见表2)。接种一定数量的细菌和真菌,间隔0天、7天、14天、21天、28天按照美国药典USP32<51>微生物防腐功效测试的检测方法检测微生物数量变化情况。同时做空白对照,将樱花叶细胞水换成去离子水。测试结果见表4。
提供实施例1的樱花叶细胞水喷雾防腐挑战测试数据,见表3。
表2:实验喷雾配方
原料 | 含量 |
丁二醇 | 2.4% |
甜菜碱 | 0.04% |
甘草酸二钾 | 0.05% |
可溶性蛋白多糖 | 0.05% |
蜂王浆提取物 | 0.05% |
樱花叶细胞水 | 余量 |
柠檬酸 | 调节PH=6~7 |
表3:实施例1樱花叶细胞水的防腐挑战测试结果数据
3、樱花叶细胞水安全性测试
HaCaT细胞为人永生表皮细胞系,对HaCaT细胞的细胞毒性,可作为对皮肤安全性的参考数据。正常细胞代谢旺盛,其线粒体内的琥珀酸脱氢酶,可将四唑盐类物质还原为带颜色的结晶状物质,沉积在细胞周围,该变化可通过酶标仪读取OD值,通过OD值与空白对照组的比较,可以得知细胞的相对生长情况。提供实施例1樱花叶细胞水的安全性测试结果,见附图1。
由图1数据说明了樱花叶细胞水对人体表皮细胞基本无毒性,安全性高。
实施例和对比例工艺结果分析:
表4:实施例和对比例所提取樱花叶细胞水的主要挥发性活性成分含量表及防腐挑战测试结果
续表4:
通过实施例和对比例比较,本发明先通过低温-真空(减压)技术对樱花叶进行初步提取,再利用初提细胞液的高渗透性,将其作为酶解溶剂,倒回初提残渣中并再次低温-真空(减压)提取得到的樱花叶细胞水,挥发性活性组分多且含量高。具体的,对比例1采用单一的低温真空提取技术,提取率较低,且成分较少,香味淡。由对比例2可知,单一的酶提取法得到的细胞水含有较多杂质,异味重,防腐性差;而且需额外加入水作为酶解溶剂,稀释了提取液,同时提取的挥发性活性成分少,导致香味淡,应用价值不高。对比例3先对樱花叶进行酶解一段时间,再采用低温-真空(减压)技术提取,会有酶解过度产生杂味的问题;而且采用水作为酶解溶剂,由于水的渗透性差,且外加的水会稀释提取液,导致了提取率较低、细胞水香味淡等缺陷。由对比例4可知,由于初提时间过短,初提细胞水太少,导致酶解程度过低,相对于实施例1提取率较低,并且活性成分含量少导致香味较淡。
Claims (8)
1.一种樱花叶细胞水的提取方法,其特征在于,包括以下步骤:不加入溶剂,将樱花叶在温度30℃~65℃、压力-60kPa~-101kPa下初步提取,樱花叶细胞水形成蒸气并冷凝收集液体,提取1.5~2.5小时,得到初提细胞水和初提樱花叶残渣,在初提细胞水中加入初始樱花叶总重量为基准的0.25%~0.45%的纤维素酶和0~0.1%的果胶酶后再倒回初提樱花残渣中,在温度35℃~50℃、压力-85kPa~-101kPa条件下再次提取5~7小时,收集得到樱花叶细胞水。
2.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,所述的初步提取温度为35℃~50℃、压力条件为-85kPa~-101kPa。
3.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,纤维素酶的加入量为0.3%~0.4%,果胶酶加入量为0.01%~0.06%。
4.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,在提取过程中进行搅拌,搅拌速度为1~150转/分钟。
5.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,收集过程中进行冷凝,冷凝温度为-10℃~8℃。
6.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,所述的樱花叶切割成宽度为1mm~10mm。
7.根据权利要求1所述的樱花叶细胞水的提取方法,其特征在于,所述的樱花叶无霉变且未腐烂。
8.权利要求1~7任一项所述的提取方法得到的樱花叶细胞水的应用,其特征在于,用于食品、保健品、药品及化妆品领域。
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CN207575846U (zh) * | 2017-12-05 | 2018-07-06 | 天津芸熙生物技术有限公司 | 一种用于樱花叶微波连续提取设备 |
CN109730948A (zh) * | 2019-01-24 | 2019-05-10 | 山东贝世康生物科技有限公司 | 从牡丹鲜花中提取牡丹鲜花细胞水的方法及应用 |
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