CN102883627B - 将酵母接种到果汁中的方法 - Google Patents
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- C12G1/0203—Preparation of must from grapes; Must treatment and fermentation by microbiological or enzymatic treatment
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Abstract
本发明提供冷冻形式的新型葡萄酒酵母产品。该产品在发酵罐中生产,浓缩,加入冷冻保护剂。然后在-50°C下冷冻这种混合物。这种产品的特征在于,除其是冷冻的事实外,其可直接添加到葡萄汁中而不要求复水,因为酵母在生产过程中没有脱水。
Description
发明领域
本发明涉及将酵母接种到果汁或水果中的方法。总的来说,这对饮料工业,尤其是对酿酒工业而言是重要的。
背景技术
最近50年,酿酒师已习惯在酒中接种纯酵母培养物。这样的接种培养物也可以被称作“起子培养物”。酵母是在发酵罐中生产、浓缩并随后在转鼓上干燥(所谓流化床干燥工艺)的被称作活性干酵母(ADY)的脱水形式。其在本文中也被称作冻干酵母。
由于酵母脱水的事实,其因此在施用于底物,如酿酒情况中的葡萄汁之前需要复水才能代谢活跃。这对酵母细胞而言是非常精妙的过程并需要非常注意和小心确保酵母细胞以使细胞可存活的方式水合。
如Roger Boulton等人1996年的教科书的第124页,左栏中论述,在活性干酵母生产的干燥过程中,酵母细胞的细胞膜丧失它们的渗透屏障功能。因此,为了重建这种功能,重要的是通过将酵母添加到40°C水中20分钟来使该膜复水。因此,酵母复水过程通常涉及在35-38°C的未氯化水或水/葡萄汁混合物(2:1)中复水20-30分钟,接着添加相同体积的葡萄汁(50:50果汁/水掺合物),其在添加到酒中之前保持另外20-30分钟。重要的是,该葡萄汁不应含有任何SO2,其会在敏感的复水过程中杀灭酵母细胞(O’Kennedy,2008)。此外,该复水混合物必须在20分钟后用在水中的果汁冷却,每次5°C。在30分钟后无法从复水温度冷却也会造成显著的细胞死亡(O’Kennedy,2008)。此外,应小心使用未污染的葡萄汁进行复水程序,因为用污染的葡萄汁复水会造成使用该复水混合物接种的整个葡萄酒发酵的污染。不同制造商提出对这种程序的变动,但关键步骤在于脱水细胞需要在特定温度、条件下在水或水/果汁混合物中暴露特定时间以适当水合,由此避免细胞死亡和随后失活。不推荐将活性干酵母直接添加到酒中,因为葡萄汁中的高糖浓度、SO2和其它化合物不允许酵母最佳复水。因此,葡萄酒酵母制造商无一提出将活性干酵母直接接种到葡萄汁(grape must)中。
如技术人员已知的是-在本文中,术语果汁(fruit juice)和果汁(fruit must)被理解为相同,这些术语在本文中可互换使用。
如Soubeyrand等人2006中所述,活性干酵母(冻干)在没有最佳复水时丧失活性。活性干酵母的不恰当复水也会造成酒精发酵停滞(O’Kennedy,2008)。因此,活性干酵母的复水需要相当大量的时间和浓度并在商业酒厂在酿酒过程中通常需要大量熟练工时。
EP 1645198A1(Minaki Advance Co Ltd,Japan);EP2090647A1(Chr.Hansen);和WO2009/095137A1(Chr.Hansen)在本文中可以被视为相关现有技术。
但是,这些现有技术文献无一公开在发酵前将酵母以冷冻形式添加到果汁中的方法。
本发明涉及将酵母接种到果汁或水果中的新型方法,由此可避免上述缺点。本发明人已经发现,令人惊讶地,可以在保持酵母细胞的高存活率水平的同时将酵母的冷冻培养物(冷冻起子培养物)直接添加到葡萄汁中。
发明概述
本发明提供冷冻形式的新型葡萄酒酵母产品。该产品在发酵罐中生产,浓缩,加入冷冻保护剂。然后在合适的容器中在-50°C下冷冻这种混合物。这种产品的特征在于,除其是冷冻的事实外,其可直接添加到葡萄汁中而不要求复水,因为酵母在生产过程中没有脱水。该产品现在是含有1千克冷冻酵母细胞培养物的冷冻袋(塑料DIM袋)形式。
如本文中的实施例1中所示,当直接接种冷冻酵母时,令本发明人惊讶的是,与活性干酵母相比仅观察到细胞数的相对较小降低,而在活性干酵母中,与恰当复水相比在直接接种时观察到细胞数的大得多的降低。因此,这一特征使得冷冻酵母适合直接接种,这是一种简单的过程,其中非技术人员可以打开容器并直接将内容物添加到葡萄汁中,而不用冒着如活性干酵母的情况那样的显著细胞死亡和随之活性损失的危险。此外,酵母的直接接种防止葡萄酒发酵可能被污染的复水混合物破坏。
因此,根据这种令人惊讶的发现,可随之理解,根据本发明的冷冻酵母的使用非常有利于葡萄酒生产。
另一优点在于,直接接种显著节省酿酒师的时间和人力,因为活性干酵母的复水花费1小时并需要技术人员制备高度特异的复水底物,而冷冻酵母的直接接种花费数分钟并且不需要专业训练或技术,简单地将该产品添加到葡萄汁或压榨葡萄中。
已使用葡萄酒酵母Pichia kluyveri获得本文中提供的结果。技术人员可容易地将该技术转用于其它类型的酵母,如酿酒酵母、巴斯德酵母、贝酵母或Torulaspora delbreuckii、耐热克鲁维酵母等。
该产品也可用于制造葡萄酒以外的饮料,如啤酒、苹果汁、清酒(sake)、酸乳酒(kefir)、软饮料和其中需要特定酵母的作用并以方便的直接接种形式施用的饮料。该起子培养物可用于发酵任何类型的糖基培养基,如葡萄汁或苹果汁或全果。
附图
图1和2:这些图在此显示如本文中的实施例1中详细论述的相关实验结果。
详述
一方面,本发明涉及将酵母接种到果汁或水果中的方法。所述酵母造成发酵,这种发酵的产物可用于食品工业,更特别是饮料工业。所设想的饮料是例如葡萄酒、啤酒、苹果汁或通过发酵水果或植物材料,如果汁、蔬菜、植物或水果产生的其它饮料。水果或植物材料的实例是葡萄、葡萄汁、苹果或苹果汁或大麦。
所用酵母可以是饮料或食品工业中使用的任何类型的酵母。实例包括例如Pichia kluyveri、酿酒酵母、巴斯德酵母、贝酵母、Torulasporadelbreuckii或耐热克鲁维酵母等。
在酿酒工业中普遍使用酵母。葡萄酒生产中所用的典型酵母来自酵母科(子囊菌酵母)。常使用来自酵母属的酵母[例如物种酿酒酵母(SC)]。另一些使用的酵母来自相同的酵母科但来自其它属,如克鲁维酵母菌属[例如物种耐热克鲁维酵母(KT)]和有孢圆酵母属[例如物种戴尔有孢圆酵母(TD)]。
为了接种发酵水果或植物材料,可以使用纯酵母培养物(即仅含一种类型的酵母的培养物),但也可以使用两种或更多种类型的酵母的混合培养物作为接种体。
对混合起子培养物而言,知道最终市售起子培养物中各酵母物种的特定量/比率是重要的。这对包含酵母属、克鲁维酵母菌属和/或有孢圆酵母属的葡萄酒起子培养物而言特别如此。
可以通过发酵酵母、接着离心收取酵母来制造起子培养物。这产生液体酵母生物质。通常,1000升发酵罐产生大约100千克液体糊形式的“湿重酵母”。基于湿重,这种糊通常含有大约1010CFU(菌落形成单位)/克。在冷冻后,留下大约109CFU/克。
为了保护细胞免受严酷冷冻条件之害,可以在离心之后或之前添加冷冻保护剂。冷冻保护剂的实例包括葡萄糖、蔗糖、海藻糖。
这样的冷冻保护剂通常以大约5%-20%的量添加。该量应足以使CFU数保持在大约109或以上。
在添加冷冻保护剂和收集液体酵母生物质后,将该生物质转移到合适的容器,例如塑料袋中。这些容器随后在-10℃至-60℃的温度下冷冻。优选温度区间在-10℃和-30℃之间。
本发明的方法要求将冷冻起子培养物以足以引发和维持植物材料发酵的量添加到糖基培养基中。
因此,本发明的一个方面涉及将酵母接种到果汁中的方法,包括下列步骤:
a)在容器中提供果汁;和
b)以足以引发和维持果汁发酵的量添加冷冻形式的酵母。
技术人员在此情况下会理解,当根据上述步骤b)将冷冻形式的酵母添加到果汁中时,在此没有酵母的复水。
如技术人员理解,可以将本文中描述的本发明的方法称作直接接种方法。
在实施例1中论述了与冷冻酵母起子培养物用于接种相关的实验。令人惊讶地发现,通过在添加到葡萄汁中之前将冷冻酵母培养物解冻,葡萄汁中的细胞存活(CFU/ml)甚至好于在葡萄汁中直接接种冷冻酵母培养物时实现的细胞存活。
因此,本发明的另一方法涉及将酵母接种到果汁中的方法,包括下列步骤:
i)在容器中提供果汁;
ii)提供冷冻形式的酵母;
iii)将该冷冻酵母解冻以获得液体形式的酵母;和
iv)以足以引发和维持果汁发酵的量添加液体形式的酵母。
技术人员在此情况下会理解,当根据上述步骤iii)解冻冷冻酵母以获得液体形式的酵母和随后将液体形式的酵母添加到果汁中时,在此没有酵母的复水。
如技术人员理解,酿酒厂中的果汁发酵步骤的温度通常为15至25°C。因此,当酿酒厂中的人将冷冻形式的酵母添加到果汁中[即根据上述步骤b)]时,该酵母冷冻形式通常在15至25°C的温度下解冻。
在本文中的实施例1中,解冻温度为30°C。
不受制于理论,但相信,通过上述步骤iii)的解冻温度为25至35°C,获得本文中的最佳结果。
如技术人员理解,可以例如通过将冷冻形式的酵母添加到例如水或水/果汁混合物中来进行上述步骤iii)的解冻,其中该水或水/果汁混合物具有合适的温度。如技术人员理解,由此获得液体形式的酵母,其可根据上述步骤iv)添加到果汁中。
实施例
材料和方法
此实施例中使用的活性干酵母是贝酵母B52(Lesaffre)。此实施例中使用的冷冻酵母是Pichia kluyveri(Chr.Hansen)。所有接种一式三份进行,在下图1和2中的柱形图中指出标准误差。在用酵母接种后,将该混合物充分搅拌以溶解内容物,然后进行标准稀释以便平板接种。为了测定这些产品的细胞数,在蛋白胨水中进行稀释并将1毫升倒入陪替氏培养皿中,此后在40°C下倒入液体形式的酵母葡萄糖培养基(YGM)琼脂。将板在30°C下培养3天,此后计数菌落数。基于ADY和冷冻酵母的原始重量添加计算如下图1和2中所示的CFU/g。
为了测定在合成葡萄汁培养基中接种后的酵母细胞计数,遵循相同程序。将50克/升的量的冷冻或解冻的Pichia kluyveri酵母接种到在室温(21°C)下的200毫升合成葡萄汁培养基中。
将活性干酵母复水(20分钟37°C,1克在10毫升未氯化水中)或称出以用0.3克/升进行直接接种,相当于2*106CFU/毫升。对直接接种和复水的ADY而言,合成葡萄汁(grape must)的温度必须调节至30°C以由于接种体与培养基的温度差而利于更大存活。
根据Costello等人2003以250克/升的糖浓度和3.5的pH制备合成葡萄汁(grape must)。在接种后,在蛋白胨水中进行稀释并将1毫升倒入陪替氏培养皿中,此后在40°C下倒入液体形式的酵母葡萄糖培养基(YGM)琼脂。将板在30°C下培养3天,此后计数菌落数。使用添加到合成葡萄汁(grape must)中的酵母物料的量和由板计数获得的CFU/毫升值,可以计算以CFU/克计的值。
实施例1:冷冻酵母的使用-与冻干酵母比较
在图1中显示与先在37°C水中复水ADY酵母20分钟和随后在合成葡萄汁(grape must)中接种(垂直线)相比,在合成葡萄汁中直接接种时(水平线)活性干酵母(ADY)的直接接种的有害作用的结果。在接种后,通过倾注平板法和用于计算平均CFU/克值的数据进行细胞计数。原始活性干酵母的CFU/克平均计算为2.40E+10(虚线)。活性干酵母的直接接种得出4.49E+9的平均CFU/克计算值,意味着81.3%的细胞由于直接接种而死亡。但是,当先在37°C水中复水20分钟和随后在合成葡萄汁中接种时,平均CFU/克计算值为2.35E+10,意味着在复水和接种后只有大约2.1%的细胞死亡。
在图2中显示在先解冻和不解冻的情况下在合成葡萄汁(grapemust)中直接接种冷冻酵母的效果。原始冷冻酵母产品的细胞计数为8.0E+8(虚线)。在葡萄汁中直接接种冷冻酵母(无解冻)后,细胞计数平均计算为5.1E+8(水平线),在解冻和随后直接接种在葡萄汁中后,细胞计数平均计算为7.1E+8(垂直线),意味着在冷冻直接接种(无解冻)中,只有36.3%的细胞死亡,在冷冻直接接种(解冻后)中,只有11.3%的细胞死亡。这与活性干酵母相比非常有利,其中81.3%的细胞死亡,意味着冷冻酵母可用于直接接种,而没有细胞计数和随后活性的显著损失。
结论:
这种实施例1的结果表明,活性干酵母(冻干)在直接接种时(无复水)与在水中复水(特定温度)和果汁稀释后接种相比损失显著量的活细胞-即这些结果符合如上论述的现有技术知识。
但是,当直接接种冷冻酵母时,令本发明人惊讶的是,如在上述情况中那样仅观察到细胞数的相对较小降低。
因此,根据这种令人惊讶的发现,可随之理解,根据本发明的冷冻酵母的使用非常有利于葡萄酒生产。
另一优点在于,直接接种显著节省酿酒师的时间,因为活性干酵母的复水花费多于1小时,而冷冻酵母的直接接种花费数分钟。
如上述实施例中所示,令人惊讶地发现,通过首先将冷冻培养物解冻和随后直接接种在葡萄汁中,细胞存活(CFU/ml)甚至好于没有先解冻就在葡萄汁中直接接种冷冻酵母培养物时实现的细胞存活。
参考文献:
1.EP 1645198
2.EP 2090647
3.WO 2009/095137
4.Peter Costello,Paul Henschke和Andrew Markides(2003)Standardised methodology for testing malolactic bacteria and wine yeastcompatibility.Australian Journal of Grape and Wine Research 9(2):127–137.
5.Roger Boulton,Vernon Singleton,Linda Bisson和Ralph Kunkee.1996.Principles and Practices of Winemaking.pp.124.
6.Karien O’Kennedy.(2008).How to avoid stuck fermentations,The Australian&New Zealand Grapegrower&Winemaker.November,Issue 538,103-105.
7.Virginie Soubeyrand,Anne Julien和Jean-Marie Sablayrolles(2006)Rehydration Protocols for Active Dry Wine Yeasts and the Searchfor Early Indicators of Yeast Activity American Journal of Enolology andViticulture.57(4)474-480.
Claims (10)
1.生产饮料的方法,包括下列步骤:
a)提供水果介质
b)加入酵母发酵上述水果介质,
其中,酵母以冷冻形式加入或在加入水果介质之前,冷冻酵母解冻以获得液体形式的酵母,
其中饮料选自葡萄酒、啤酒、苹果汁和软饮料。
2.生产饮料的方法,包括下列步骤:
a)提供蔬菜介质
b)加入酵母发酵上述蔬菜介质,
其中,酵母以冷冻形式加入或在加入蔬菜介质之前,冷冻酵母解冻以获得液体形式的酵母,
其中饮料选自葡萄酒、啤酒、清酒和软饮料。
3.按照前述权利要求1的方法,其中水果介质是水果汁。
4.按照前述权利要求1的方法,其中水果介质是葡萄汁,并且饮料是葡萄酒。
5.按照前述权利要求1或2的方法,其中酵母选自酵母属、克鲁维酵母菌属和有孢圆酵母属。
6.按照权利要求5的方法,其中酵母选自Pichia kluyveri、酿酒酵母、巴斯德酵母、贝酵母、Torulaspora delbreuckii或耐热克鲁维酵母。
7.按照前述权利要求1或2的方法,其中冷冻形式的酵母包含冷冻保护剂。
8.按照权利要求7的方法,其中冷冻保护剂以5%-20%的数量存在。
9.按照前述权利要求1或2的方法,其中冷冻酵母包含至少109CFU/克。
10.按照前述权利要求1或2的方法,其中通过发酵酵母、离心收取酵母来提供液体酵母生物质以及冷冻液体酵母生物质来获得酵母。
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CA3107379A1 (en) | 2018-08-13 | 2020-02-20 | Chr. Hansen A/S | Production of alcohol-free fermented vegetable juice with pichia kluyveri yeast |
WO2020070286A1 (en) | 2018-10-04 | 2020-04-09 | Chr. Hansen A/S | Yeast for beer production |
CN109486577A (zh) * | 2018-12-29 | 2019-03-19 | 大连民族大学 | 一种果汁发酵的甜酒酿 |
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JP6196651B2 (ja) | 2017-09-13 |
JP2016039816A (ja) | 2016-03-24 |
WO2011134952A1 (en) | 2011-11-03 |
MX2012012522A (es) | 2012-12-17 |
US20130045301A1 (en) | 2013-02-21 |
AU2011247632B2 (en) | 2014-10-02 |
EP3248471A1 (en) | 2017-11-29 |
JP6096110B2 (ja) | 2017-03-15 |
EP2563169A1 (en) | 2013-03-06 |
RU2012150430A (ru) | 2014-06-10 |
BR112012027549A2 (pt) | 2015-09-15 |
MX345378B (es) | 2017-01-27 |
IL222658A0 (en) | 2012-12-31 |
JP2013524827A (ja) | 2013-06-20 |
US11311032B2 (en) | 2022-04-26 |
ES2637952T3 (es) | 2017-10-18 |
BR112012027549A8 (pt) | 2015-10-06 |
AU2011247632A1 (en) | 2012-11-22 |
EP2563169B1 (en) | 2017-06-14 |
NZ603267A (en) | 2014-12-24 |
CL2012003021A1 (es) | 2012-11-30 |
CA2797597A1 (en) | 2011-11-03 |
PL2563169T3 (pl) | 2017-11-30 |
CN102883627A (zh) | 2013-01-16 |
DK2563169T3 (en) | 2017-09-11 |
CA2797597C (en) | 2020-12-08 |
BR112012027549B1 (pt) | 2021-08-17 |
RU2585846C2 (ru) | 2016-06-10 |
HUE033635T2 (en) | 2017-12-28 |
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