CN114958652B - 一种含植物乳杆菌y44的复配发酵剂及其发酵复配乳 - Google Patents
一种含植物乳杆菌y44的复配发酵剂及其发酵复配乳 Download PDFInfo
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Abstract
本发明公开了一种含植物乳杆菌Y44的复配发酵剂及复配乳与应用,属于微生物技术领域。一种植物乳杆菌(Lactobacillus plantarum)Y44作为发酵剂的应用,所述植物乳杆菌(Lactobacillus plantarum)Y44于2018年8月21日,保藏于中国典型培养物保藏中心,菌株保藏号为CCTCC NO:M 2018558。本发明提供一种脱脂乳与豆乳复配的发酵乳,通过多人感官评定筛选出比例为豆乳:脱脂乳=2:8,此比例具有较好的理化性质和ACE抑制率,整体口感较好,除传统的奶香味,还具有独特的豆香味以及特殊风味物质。复配比例为2:8的时候,整体物理表征比较好,多项指标均为较佳,综合评定为最佳比例。该复配发酵乳具有较好理化指标和降血压功能。本发明所述的复配乳具有降低高血压的作用,具有很好的应用前景。
Description
技术领域
本发明涉及一种含植物乳杆菌Y44的复配发酵剂及其发酵复配乳,属于微生物技术领域。
背景技术
酸奶是一种传统发酵乳制品,具有丰富的营养价值和良好的保健功能。具有易于消化和吸收,能促进胃液分泌,提高食欲,较高的营养及保健功能,因此得到了消费者的青睐。酸奶发酵剂是生产酸奶、保证其质量稳定及形成酸奶优良的感官品质以及组织状态的关键所在。发酵剂性能决定了酸奶发酵周期、酸奶理化特性。而生产酸奶的原料乳及配料也是决定酸奶营养功能的重要因素。
目前,我国酸奶产量不断增加,品种日益丰富。而国内发酵生产酸奶所使用的发酵剂菌种大多为嗜热链球菌和保加利亚乳杆菌,产品质量较稳定、生产周期较短、成本低。近几年,益生菌发酵酸乳产品种类日益增多且深受消费者喜爱。益生菌是指一类非致病的经过消化系统对人健康产生积极作用的微生物,它们能够改善人体肠道菌群结构从而能维持其健康,益生菌相关产品深受关注。故采用益生菌与商业酸奶发酵剂进行复配,以获得具有益生功能的复配发酵剂。
大豆产品在亚洲国家消费量较高。然而,消费者不接受豆制品中的豆腥味和青草味,同时大豆中的低聚糖可能会引起胃肠胀气,因此限制豆制品在西方国家的消费。与牛乳类似,豆乳中含有低聚糖和氨基酸,可以维持益生菌的生长。有研究表明,利用益生菌发酵豆乳可以解决大豆的风味不良和肠胃胀气的问题,但是也有研究表明发酵生产的大豆酸奶存在钙含量低、风味差、质地粗糙等缺点。另外,豆乳不能维持某些微生物的生长,例如保加利亚乳杆菌,从而导致产酸不足。
发明内容
为解决现有技术存在的问题,本发明将豆乳与牛乳混合,以植物乳杆菌Y44、保加利亚乳杆菌6047和嗜热链球菌6038进行混合作为发酵剂,获得营养、风味和质地优良的脱脂乳豆乳复配发酵乳。发现豆乳和脱脂乳以2:8复配混合制备的酸奶的品质最好。
本发明提供了一种植物乳杆菌(Lactobacillus plantarum)Y44作为发酵剂的应用,所述植物乳杆菌(Lactobacillus plantarum)Y44于2018年8月21日,保藏于中国典型培养物保藏中心,菌株保藏号为CCTCC NO:M 2018558。
本发明还提供了一种复配发酵剂,所述发酵剂的体积比例范围为1:1:1~1:1:10,优选体积比为1:1:1的植物乳杆菌(Lactobacillus plantarum)Y44、商业酸奶发酵剂保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcusthermophilus)6038组成。
本发明还提供了一种复配发酵乳,将所述的复配发酵剂接种于复配乳中制备而成。
进一步地,上述技术方案中,所述复配乳为脱脂乳和豆乳,所述脱脂乳和豆乳的体积比为1~17:0~3。
所述脱脂乳和豆乳的体积比优选为8:2。
进一步地,上述技术方案中,所述脱脂乳的总乳固体浓度为11.5%~12%(w/v),优选为12%(w/v);所述豆乳为大豆与水以1:6~1:7的比例混合然后磨制过滤而成,优选为1:7。
进一步地,上述技术方案中,升水复配乳中包括糖,所述糖包括白砂糖,升水白砂糖在复配乳中的含量为4%~6%(w/v),优选为6%(w/v)。
进一步地,上述技术方案中,将复配发酵剂接种于复配乳中制备复配发酵乳,所述复配发酵剂在复配乳中的接种量为3%~5%(v/v),优选为4%(v/v);发酵的条件为40℃~42℃,优选为42℃的条件下发酵至凝乳,然后4~8℃后熟10~12h,优选为4℃后熟12h。
进一步地,上述技术方案中,所述复配发酵乳的活菌数为4.4-4.7×109cfu/ml,pH值是4.55-4.65,滴定酸度为85-89°T。
本发明还提供了所述的复配发酵乳在缓解高血压或者降低高血压中的应用。
本发明所提供的复配发酵乳还具有良好的抗氧化活性、良好的胃、肠转运耐受性、调节肠道微生物群来改善肠道炎症和脂质代谢紊乱的作用。
本发明将植物乳杆菌与商业酸奶发酵剂制备复配发酵剂,并制备豆乳和脱脂乳的不同比例的复配乳,利用复配发酵剂发酵复配乳,所获得的复配发酵乳具有良好的理化特性,并具有缓解高血压的应用,属于发酵乳制品技术领域。本发明提供一种脱脂乳与豆乳复配的发酵乳,通过多人感官评定筛选出比例为豆乳:脱脂乳=2:8,此比例具有较好的理化性质和ACE抑制率,整体口感较好,除传统的奶香味,还具有独特的豆香味以及特殊风味物质。复配比例为2:8的时候,整体物理表征比较好,多项指标均为较佳,综合评定为最佳比例。该复配发酵乳具有较好理化指标和降血压功能。
本发明所述的植物乳杆菌(Lactobacillus plantarum)Y44发酵的复合乳具有降低高血压的作用,具有很好的应用前景。
附图说明
图1为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的硬度。
图2为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的滴定酸度。
图3为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的持水力。
图4为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的游离氨基酸含量。
图5为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的双乙酰含量。
图6为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的感官评价得分。
图7为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳的ACE抑制活性。
图8为植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillus bulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038复配发酵乳对高血压SHR大鼠降血压效果。
具体实施方式
下述非限定性实施例可以使本领域的普通技术人员更全面地理解本发明,但不以任何方式限制本发明。
实施例1菌株培养
本发明的植物乳杆菌Y44、保加利亚乳杆菌6047和嗜热链球菌6038在MRS培养基中进行培养活化,具体步骤如下:
菌株培养方法:
(1)MRS培养基制备
按照20g/L葡萄糖,10g/L蛋白胨,5g/L酵母膏,10g/L牛肉膏,1mL吐温-80,2g/L磷酸氢二钾,2g/L柠檬酸二铵,5g/L乙酸钠,0.58g/L七水硫酸镁,0.25g/L四水硫酸锰,称取这些原料,混合均匀,将得到的混合物混溶于去离子水中,得到的溶液在温度121℃下灭菌20min,得到所述的MRS培养基。
(2)菌株的培养
植物乳杆菌(Lactobacillus plantarum)Y44(于2018年8月21日,保藏于中国典型培养物保藏中心,菌株保藏号为CCTCC NO:M 2018558,序列如SEQ ID NO.1所示)、保加利亚乳杆菌(Lactobacillus bulgaricus)6047(CICC 6047,中国工业微生物菌种保藏管理中心)和嗜热链球菌(Strepococcus thermophilus)6038(CICC 6038,中国工业微生物菌种保藏管理中心)划线接种于含有2%琼脂的步骤(1)培养基中,在温度37℃下培养48h~72h,挑取单菌落,接种到MRS液体培养基中在37℃下培养18h。
实施例2菌液的制备
取植物乳杆菌(Lactobacillus plantarum)Y44、保加利亚乳杆菌(Lactobacillusbulgaricus)6047和嗜热链球菌(Strepococcus thermophilus)6038(以下用Y44、6047和6038表示)分别接种于MRS培养基中活化3次。活化后的菌株以2%(v/v)接种量于MRS培养液,37℃培养18h。将Y44、6047和6038的MRS培养液离心(5000rmp,5min)获得菌体沉淀物,用0.85%生理盐水清洗3次,重悬至0.85%生理盐水中,调整Y44、6038和6047的菌悬液浓度为3×108CFU/mL。
实施例3复配发酵乳的制备
将市售黄豆,快速用超纯水清洗2~3次,洗去灰尘,然后加入2倍豆重的超纯水浸泡过夜。在浸泡过夜后的黄豆中加入7倍豆重的超纯水放入豆浆机中进行磨浆,磨浆5~10min后,将豆乳用双层滤布过滤,去除滤渣。过滤液即为豆乳,冷却后备用。称量一定数量的复原脱脂乳粉,按照脱脂乳粉:水=12:88的比例加入水,用玻璃棒将其搅拌均匀,获得复原脱脂乳。按照以下比例(体积比)对脱脂乳和豆乳进行混合获得复配乳:第一组(豆乳:脱脂乳=0:1),第2组(豆乳:脱脂乳=1:9),第3组(豆乳:脱脂乳=3:17),第4组(豆乳:脱脂乳=2:8)。脱脂乳和豆乳按照上述比例混合后进行灭菌处理(105℃,15min),冷却后备用。
Y44、6047和6038菌株分别在生理盐水中制备菌悬液,Y44、6047和6038的菌悬液按1:1:1(体积比)比例混合,混合菌悬液以4%(v/v)接种量接种于不同复配乳中。在37℃条件下,在恒温培养箱里进行发酵,直至乳清析出,记录乳清析出时间;发酵结束后,4℃条件下冷藏过夜,得到复配发酵乳用于后续理化及感官指标评价。
实施例4复配发酵乳的滴定酸度
酸度用吉尔涅尔度(°T)表示。测定方法参考国标方法:GB5009.239—2016《乳和乳制品酸度的测定》。
经过测定,通过复配菌发酵的复配乳滴定酸度实验得到相应数据符合国家标准,从图2中可以看出,6038+6047+Y44(豆乳:脱脂乳=0:1)发酵乳的产酸能力强,酸度可达到112.85°T,而凝乳时间仅需6个半小时;豆乳:脱脂乳=2:8比例发酵乳的产酸能力较弱,产品的酸味没有其他组酸,品尝起来没有过酸,酸甜度适中,感官评分较好。酸偏高时乳杆菌生长速度略高于链球菌,酸奶酸度增长速度快,发酵时间缩短,利于酸奶酸度提高但不利于风味物质的形成,影响发酵乳的整体风味。随着豆乳的添加量的增加,其滴定酸度逐渐下降,整体上呈现降低趋势,整体酸度上豆乳:脱脂乳=2:8比豆乳:脱脂乳=0:1的组合要好,优选2:8的比例。
实施例5复配发酵乳的持水力
称取一定质量(W0)的复配发酵乳样品置于50mL离心管中,在4℃、3000r/min条件下离心20min后静置10min,倾倒出上清液,测定离心管内剩余样品的质量W,酸奶的持水力(WHO)计算,公式如下:WHO=W/W0×100%
通过离心称重得到复配发酵乳的持水力数据,如图3,从图中可以分析出随着豆乳添加量的增加,持水力也随之增加。整体上持水力在70%以上,豆乳:脱脂乳=2:8和0:1的组合要比1:9和3:17的组合持水能力强,差异显著。
实施例6复配发酵乳的硬度
采用TA.XT.Plus质构仪,选用P/50的圆柱型挤压检测探头,测定条件为:测前速度、测试速度为15mm/min,测后速度20mm/min,测试距离60mm,感应力Auto-4.0g。样品为4℃,储存12h的发酵乳,每个样品重复测定3次,结果取平均值。此方法为本行业通用测定发酵乳硬度方法。
对经过后熟处理的发酵乳进行质构分析得到相应数据,从图1中可以看出,不同比例复配的豆乳和脱脂乳复配乳在通过复配发酵剂(6038+6047+Y44)发酵之后相对于豆乳:脱脂乳=0:1的组合硬度差异不显著,但是硬度过大,会使发酵乳颗粒口感较硬、具有较明显的粉涩感,故优选豆乳:脱脂乳=1:9、3:17、2:8的组合。
实施例7复配发酵乳的游离氨基酸含量
采用OPA法对复配发酵乳中游离氨基酸含量进行测定,其原理是每水解一个肽键便会释放一个游离胺基,游离胺基与OPA(邻苯二苯醛)反应生成一种黄色络合物,可用分光光度计在340nm下测量其吸光度。
经过对发酵乳进行游离氨基酸实验得到相应数据,从图4中可以看出,不同比例的复配乳在通过发酵之后的发酵乳游离氨基酸含量差异显著,随着豆乳量的增加,其发酵乳的游离氨基酸含量随之增加,原因可能是大豆蛋白质含量比脱脂乳的含量高,益生菌代谢分解蛋白质数量增多,因此游离氨基酸含量明显增多。其中豆乳和脱脂乳以2:8复配的发酵乳组游离氨基酸含量显著高于其它复配组,其数值为0.304mg/mL。
实施例8复配发酵乳的双乙酰含量测定
采用邻苯二胺比色法测定复配发酵乳样品中的双乙酰的含量。以双乙酰质量浓度为横坐标,吸光值为纵坐标,绘制标准曲线。用三氯乙酸处理样品,利用紫外分光光度计在335nm处测定吸光度值,每组样品重复测三次取平均值。按照上述绘制的双乙酰标准曲线计算样品中双乙酰的含量。此方法为本行业通用测定发酵乳中双乙酰含量的方法。
对经过后熟处理的发酵乳进行实验分析得到相应数据,从图5可以看出,不同复配比例的发酵乳通过发酵之后,实验组的双乙酰含量都具有一定的含量提升,说明复配乳的不同对双乙酰的含量影响不同。随着豆乳的添加量的升高,双乙酰的含量则升高。其中豆乳:脱脂乳=2:8复配组的双乙酰含量显著高于1:0、1:9、3:17的复配组,由于双乙酰是发酵乳特殊风味的来源,这也验证了2:8复配组的风味显著高于1:0、1:9、3:17的复配组。
实施例9复配发酵乳的ACE含量
测定复配发酵样品的ACE抑制率采用紫外分光光度法,原理为ACE在37℃、pH8.3的条件下催化分解血管紧张素Ⅰ的模拟物马尿酰组胺酰亮氨酸(HHL)产生马尿酸,该物质在228nm处具有特征吸收峰。当加入ACE时,ACE对HHL的催化分解作用收到了抑制,马尿酰的生成量减少。通过测定加入抑制剂前后所生成的马尿酰紫外吸收的大小差别即可计算抑制活性的大小。此方法为本行业通用测定发酵乳中ACE抑制率的方法。
如图7所示,随着豆乳添加量的增多,实验组发酵乳的ACE抑制率在逐渐升高,说明少量的豆乳可以增加发酵乳的ACE抑制率,但是过多的添加豆乳会使ACE的抑制率降低。豆乳和脱脂乳=2:8复配的发酵乳组ACE抑制率显著高于1:0、1:9、3:17的复配组,其抑制率高达37.13%,这也说明豆乳和脱脂乳=2:8的复配组合为我们所需要的筛选组合。
实施例10复配发酵乳的感官评定
对经过后熟处理的发酵乳进行感官评定得到相应数据,从图6中可以看出,不同比例复配的豆乳和脱脂乳复配乳在通过复配发酵剂(6038+6047+Y44)发酵之后相对于纯脱脂乳风味差异显著,添加少量的豆乳可以改良产品的风味。其中豆乳和脱脂乳的复配比例为2:8时,感官评定得分高达80分。豆乳和脱脂乳的复配比例为2:8的发酵乳具有酸甜适中、良好的粘度稠度、口感顺滑、较好的质地等优点,具有独特的发酵乳气味和风味。
实施例11复配发酵乳的动物实验
将6只健康的雄性SHR(原发性高血压大鼠,购于北京维通利华)大白鼠和6只健康的雄性WKY(正常血压大鼠,购于北京维通利华)大白鼠,分为4组,每组3只,室温为23℃左右、具有通风换气设备的恒温动物房中饲养。大鼠均自由进食和饮水,实验开始前一周对大白鼠进行自由饲养以便其能够适应周围环境。
每天8:00按照10ml/kg的剂量对实验组SHR和WKY大鼠灌胃发酵乳,对照组和空白组灌胃未发酵的乳,在灌胃后3小时测定大鼠血压(BP-2010A血压计),实验进行两周。
通过分析,相比未灌胃发酵乳组,SHR-Y44发酵乳组血压降低10-15mmHg,14天后,SHR-Y44发酵乳组相对于SHR-Control组降血压效果具有显著差异,血压维持在175mmHg。WKY-Y44发酵乳组相对于WKY-Control组血压降低10-15mmHg,血压维持在135mmHg。说明复配发酵剂(6038+6047+Y44)发酵之后的豆乳和脱脂乳(2:8)复配乳具有降低血压的功能。
SEQUENCE LISTING
<110> 大连工业大学
<120> 一种含植物乳杆菌Y44的复配发酵剂及其发酵复配乳
<130> 2022
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 1546
<212> DNA
<213> 植物乳杆菌(Lactobacillus plantarum)
<400> 1
attaatttga gagtttgatc ctggctcagg acgaacgctg gcggcgtgcc taatacatgc 60
aagtcgaacg aactctggta ttgattggtg cttgcatcat gatttacatt tgagtgagtg 120
gcgaactggt gagtaacacg tgggaaacct gcccagaagc gggggataac acctggaaac 180
agatgctaat accgcataac aacttggacc gcatggtccg agtttgaaag atggcttcgg 240
ctatcacttt tggatggtcc cgcggcgtat tagctagatg gtggggtaac ggctcaccat 300
ggcaatgata cgtagccgac ctgagagggt aatcggccac attgggactg agacacggcc 360
caaactccta cgggaggcag cagtagggaa tcttccacaa tggacgaaag tctgatggag 420
caacgccgcg tgagtgaaga agggtttcgg ctcgtaaaac tctgttgtta aagaagaaca 480
tatctgagag taactgttca ggtattgacg gtatttaacc agaaagccac ggctaactac 540
gtgccagcag ccgcggtaat acgtaggtgg caagcgttgt ccggatttat tgggcgtaaa 600
gcgagcgcag gcggtttttt aagtctgatg tgaaagcctt cggctcaacc gaagaagtgc 660
atcggaaact gggaaacttg agtgcagaag aggacagtgg aactccatgt gtagcggtga 720
aatgcgtaga tatatggaag aacaccagtg gcgaaggcgg ctgtctggtc tgtaactgac 780
gctgaggctc gaaagtatgg gtagcaaaca ggattagata ccctggtagt ccataccgta 840
aacgatgaat gctaagtgtt ggagggtttc cgcccttcag tgctgcagct aacgcattaa 900
gcattccgcc tggggagtac ggccgcaagg ctgaaactca aaggaattga cgggggcccg 960
cacaagcggt ggagcatgtg gtttaattcg aagctacgcg aagaacctta ccaggtcttg 1020
acatactatg caaatctaag agattagacg ttcccttcgg ggacatggat acaggtggtg 1080
catggttgtc gtcagctcgt gtcgtgagat gttgggttaa gtcccgcaac gagcgcaacc 1140
cttattatca gttgccagca ttaagttggg cactctggtg agactgccgg tgacaaaccg 1200
gaggaaggtg gggatgacgt caaatcatca tgccccttat gacctgggct acacacgtgc 1260
tacaatggat ggtacaacga gttgcgaact cgcgagagta agctaatctc ttaaagccat 1320
tctcagttcg gattgtaggc tgcaactcgc ctacatgaag tcggaatcgc tagtaatcgc 1380
ggatcagcat gccgcggtga atacgttccc gggccttgta cacaccgccc gtcacaccat 1440
gagagtttgt aacacccaaa gtcggtgggg taacctttta ggaaccagcc gcctaaggtg 1500
ggacagatga ttagggtgaa gtcgtaacaa ggtagccgta ggagaa 1546
Claims (3)
1.一种复配发酵乳,其特征在于,将复配发酵剂接种于复配乳中制备而成;
所述的复配发酵剂由体积比为1:1:1~1:1:10的植物乳杆菌(Lactobacillus plantarum) Y44、商业酸奶发酵剂保加利亚乳杆菌(Lactobacillus bulgaricus)6047、嗜热链球菌(Strepococcus thermophilus)6038组成;
所述植物乳杆菌(Lactobacillus plantarum)Y44于2018年8月21日,保藏于中国典型培养物保藏中心,菌株保藏号为CCTCC NO:M 2018558;
所述复配乳为脱脂乳和豆乳,所述脱脂乳和豆乳的体积比为8:2;
所述脱脂乳的总乳固体浓度为11.5%~12% w/v,所述豆乳为大豆与水以1:6-1:7的比例混合然后磨制过滤而成;
将复配发酵剂接种于复配乳中制备复配发酵乳,所述复配发酵剂在复配乳中的接种量为3%~5% v/v;发酵的条件为40℃~42℃的条件下发酵至凝乳,然后4~8℃后熟10~12h;
所述复配发酵乳的活菌数为4.4-4.7×109cfu/ml,pH值是4.55-4.65,滴定酸度为85-89°T。
2. 根据权利要求1所述的复配发酵乳,其特征在于,复配乳中包括糖,所述糖包括白砂糖,白砂糖在复配乳中的含量为4%~6% w/v。
3.权利要求1-2中任一项所述的复配发酵乳在制备辅助降低血压的食品中的应用。
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