CN116832068A - 一株具有广谱抑菌活性的副干酪乳酪杆菌pc646及其应用 - Google Patents
一株具有广谱抑菌活性的副干酪乳酪杆菌pc646及其应用 Download PDFInfo
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Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
- A61K35/66—Microorganisms or materials therefrom
- A61K35/74—Bacteria
- A61K35/741—Probiotics
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Abstract
本发明提供一株分离自酸马奶的副干酪乳酪杆菌PC646,其保藏编号为CGMCC No.23515。通过胃肠液耐受性和胆盐耐受性测试确认该菌株具有良好的益生菌特性,特别是在胆盐浓度0.3%环境下培养,结果表明0.3%胆盐不会抑制该菌株的活性。此外该具备抑菌活性包括,其发酵后的无细胞提取液经脱盐、浓缩和喷雾干燥制备所得的抑菌制剂对于常见致病菌特别是金黄色葡萄球菌具有显著的抑制效果,且可以用于食品防腐保鲜。本发明为该菌株及其抑菌活性制剂的深入研究和开发利用提供了一种全新的思路。
Description
技术领域
本发明涉及微生物技术领域,具体涉及一株具有光广谱抑菌活性的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646,还涉及其在抑菌领域的应用。
背景技术
随着抗生素滥用导致细菌产生抗生素耐药性成为全球公共卫生问题,开发新的抗菌疗法成为关键,而从微生物中筛选分离天然抗菌活性物质成为解决抗生素耐药性的重要策略。其中,益生菌对致病菌的抑制特性是当前的研究热点。
益生菌抑菌功能和机理的研究结果表明,不同菌株具有不同的抑菌物质,其抑菌机理也不相同,例如有机酸通过降低微环境的pH值而增加致病菌细胞膜的通透性,导致细胞内容物泄露,从而杀死靶细菌;菌体的自聚集与致病菌的共聚集效应对抑菌活性也能起到一定的作用;此外,益生菌产生的一些非酸性抑菌物质,特别是各种细菌素,能有效抑制致病菌和食品腐败菌。因此,益生菌因其健康、安全、有效的抑菌性能而具有较高的研究价值和广阔的应用前景,将成为未来研究的主要方向。
本申请为深入研究益生菌代谢产物抑菌功能特性的开发方面提供了依据,具体的研究工作也证实了副干酪乳酪杆菌PC646代谢产物制剂在抑制致病菌方面效果确切,具有巨大的开发和利用价值。
发明内容
本发明要解决的问题是提供一株可有效抑制致病菌的副干酪乳酪杆菌,并验证该菌株在抑菌领域的应用与效果。
为了实现上述目的,本发明提供一株分离自酸马奶的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646,所述副干酪乳酪杆菌(Lacticaseibacillusparacasei)PC646于2021年9月29日保藏于中国微生物菌种保藏管理委员会普通微生物菌种保藏中心(地址:北京市朝阳区北辰西路1号院3号 ),保藏编号为CGMCC No.23515。
这株分离自中国内蒙古锡林浩特市西乌旗地区的酸马奶样品的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646,通过胃肠液耐受性和胆盐耐受性测试确认该菌株具有良好的益生菌特性,特别是在胆盐浓度0.3%环境下培养,OD620nm值增加0.3个单位所需的时间与在不含胆盐的培养基中培养的时间差为0小时,表明0.3%胆盐不会抑制该菌株的活性。因此,该菌株能够抵御消化道环境的抑制并在肠道内定植。
另一方面,本发明的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646经过感官评鉴确认具有优秀的风味和口感,且制备成褐色乳饮料后,所得产品在4℃条件下保存21天,每间隔7天测定其酸度和益生菌活菌数,确认其性质变化较小,在贮藏期间始终保持高活菌数,证明本发明的PC646具有良好的益生特性,可应用于商业化发酵乳制品中。
本发明提供上述副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646在抑制所述致病菌是肠沙门氏菌(Salmonella enterica)、荧光假单胞菌(Pseudomonasfluorescens)、铜绿假单胞菌(Pseudomonas aeruginosa)、伴放线放线杆菌(Actinobacillus actinomycetemcomitans)或哈维氏弧菌(Vibrio harveyi);在革兰氏阳性菌中,对金黄色葡萄球菌(Staphylococcus aureus)、单增李斯特氏菌(Listeriamonocytogenes)、蜡样芽孢杆菌(Bacillus cereus)或变异链球菌(Streptococcusmutans)中的应用。
本发明还提供副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646在食品添加剂的应用,特别地,所述食品添加剂是食品防腐剂。
本发明还提供一种具有抑菌特性的益生菌粉剂的制备方法,所述方法包括以下步骤:
(1)种子培养
将冷冻保存在-80℃的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646接种至MRS液体培养基中,于37℃培养24h,以体积分数2%的接种量连续活化至第3代,培养18h后,得到种子液;
(2)扩大培养
将所述种子液以体积比5%的接种量接入强化MRS液体培养基中(含有40g/L葡萄糖的MRS液体培养基),然后加入氨水或氢氧化钠调整发酵液初始pH至6.50±0.02,空气保压0.02~0.03MPa,设置搅拌速度100rpm/min,再温度37±0.2℃条件下进行发酵,使用氨水作维持pH恒定(5.50-6.00)±0.02;
(3)发酵液的后处理
将步骤(2)发酵至终点的发酵液高速离心去除菌体,收集到的无细胞提取液经脱盐、浓缩和喷雾干燥制备得到粉剂。
本发明通过实验验证了副干酪乳酪杆菌PC646具备抑菌活性包括:
其发酵后的无细胞提取液经脱盐、浓缩、喷雾干燥制备所得的粉剂可有效抑制荧光假单胞菌、铜绿假单胞菌、蜡样芽孢杆菌、单增李斯特菌、变异链球菌;
其发酵后的无细胞提取液经脱盐、浓缩、喷雾干燥制备所得的粉剂可延缓肠炎沙门氏菌的生长;
其发酵后的无细胞提取液经脱盐、浓缩、喷雾干燥制备所得的粉剂经40倍稀释(约2.5%干物质)仍对金黄色葡萄球菌具有显著的抑制作用。
其发酵后的无细胞提取液经脱盐、浓缩、喷雾干燥制备所得的粉剂在熟牛肉防腐中的应用。
因此,该菌株具有较强的抑菌潜力,其发酵后的无细胞提取液经脱盐、浓缩、喷雾干燥制备所得的粉剂对于致病菌具有显著的抑制效果。
本发明涉及以下微生物:
副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646,于2021年9月29保藏于中国微生物菌种保藏管理委员会普通微生物菌种保藏中心(地址:北京市朝阳区北辰西路1号院3号 ),保藏号为CGMCC No.23515。
附图说明
图1副干酪乳酪杆菌PC646代谢产物制剂对金黄色葡萄球菌的抑制作用;
图2不同蛋白酶处理对副干酪乳酪杆菌PC646代谢产物制剂的抑菌活性影响;
图3副干酪乳酪杆菌PC646代谢产物制剂对熟牛肉金黄色葡萄球菌抑制效果;
图4副干酪乳酪杆菌PC646代谢产物制剂对熟牛肉酸碱度的影响;
图5副干酪乳酪杆菌PC646代谢产物制剂对熟牛肉色度的影响;
图6副干酪乳酪杆菌PC646代谢产物制剂对熟牛肉挥发性盐基氮的影响;
图7副干酪乳酪杆菌PC646代谢产物制剂对熟牛肉硫代巴比妥酸的影响。
具体实施方式
下面将结合实施例对本发明作进一步说明,以下实施例仅为说明性的,本发明并不受这些实例的限制。
在本发明中,如无特殊说明,用于解释浓度的“%”均为重量百分比。
本发明涉及以下培养基:
MRS液体培养基:大豆蛋白胨10g、牛肉浸粉8g、酵母浸粉4g、葡萄糖20g、Tween-801ml、磷酸氢二钾2g、乙酸钠5g、柠檬酸三胺2g、硫酸锰0.05g、硫酸镁0.2g,蒸馏水1L,调节pH至6.5±0.2,121℃,15min灭菌。
MRS固体培养基:大豆蛋白胨10.0g、牛肉浸粉8.0g、酵母浸粉4.0g、葡萄糖20.0g、Tween-80 1.0ml、磷酸氢二钾2.0g、乙酸钠5.0g、柠檬酸三胺2.0g、硫酸锰0.05g、硫酸镁0.2g,蒸馏水1L,调节pH至6.5±0.2,琼脂15g,121℃、15min灭菌。
LB培养基:胰蛋白胨10.0g、酵母浸粉5.0g、氯化钠10.0g,7.0±0.1,琼脂18g,121℃、15min灭菌。
NA培养基:蛋白胨10.0g、牛肉粉3.0g、氯化钠5.0g、蒸馏水1L,调节pH至7.3±0.1,琼脂15g,121℃、15min灭菌。
本发明涉及以下测定方法:
酸碱度测定:将样品(1g)和9mL蒸馏水均质1min,使样品与蒸馏水充分混匀,用便携式pH计测定匀浆的pH值。
色度测定:使用色度计,测量参数包括L*(亮度)、a*(发红度)和b*(黄度)。此外,总色差(ΔE*)由以下公式计算:ΔE*=[(ΔL*)2+(Δa*)2+(Δb*)2]1/2。其中ΔL*、Δa*和Δb*是样品组和未处理的熟牛肉的L*、a*和b*的差异。
挥发性盐基氮(TVB-N)测定:参照GB 5009.228-2016《食品安全国家标准食品中挥发性盐基氮的测定》。
硫代巴比妥酸反应物(TBARS)测定:参照GB/T 35252-2017《动植物油脂2-硫代巴比妥酸值的测定直接法》。
实施例1副干酪乳酪杆菌PC646的分离与鉴定
将从内蒙古锡林浩特市西乌旗地区的酸马奶样品中采集到的样品倍比稀释,吸取10-5和10-6稀释梯度样品200μL均匀涂布于MRS固体培养基平皿内,涂布均匀后于37℃进行厌氧培养(48±2)h。挑取形态、大小、颜色不同的单个菌落接种于液体培养基中于37℃恒温培养箱培养(24±0.5)h。待菌株生长良好后,进行革兰氏染色、镜检,保存分离株。
本菌株显微镜下观察形态为革兰氏阳性菌,菌体呈短杆状;MRS固体培养基上菌落直径2-3毫米,菌落呈乳白色或白色,且质地柔软。
对分离到的菌株进行生理生化鉴定和16S rRNA分子鉴定,分子序列经NCBI数据库Blast比对,确认PC646为副干酪乳酪杆菌(Lacticaseibacillus paracasei),其16S rRNA序列如SEQ ID No.1所示。
实施例2副干酪乳酪杆菌PC646代谢产物的抑菌活性测试
将冷冻保存在-80℃的菌株接种至MRS液体培养基中,于37℃培养24h,以体积分数2%的接种量连续活化至第3代,培养18h后,将发酵液以6000×g离心10min,弃除菌体,将所得无细胞提取液在50℃下用旋转蒸发仪浓缩5倍,即得到无细胞浓缩液,将pH调至7.0,以E.coli CICC 23657,S.aureus ATCC 12600为指示菌,对25种指示菌进行抑菌实验,包括10种革兰氏阴性菌、12种革兰氏阳性菌(其中6种益生菌)、3种真菌应用抑菌圈法筛选产抑菌活性物质的乳酸菌。
将指示菌从体积分数15%的甘油冻存管中接种至LB培养基中,于37℃、200r/min的摇床中培养12h,用PBS磷酸盐缓冲液(0.01mol/L,pH 7.3)将指示菌稀释成105CFU/mL的菌悬液。将配置好的NA培养基高温灭菌后冷却至45℃,将稀释好的指示菌菌悬液按照体积分数2%加入NA培养基,充分搅拌均匀后量取20mL倒入灭菌平皿,凝固后放置直接8mm牛津杯。用移液枪吸取150μL样品加入牛津杯,平皿转移至冰箱4℃扩散4h,然后置于37℃培养箱培养10-12h,测量并记录抑菌圈。
将上述种子液以5%(V/V)的接种量进行扩大培养。接种结束后加入氨水或氢氧化钠调整发酵液初始pH至6.50±0.02,空气保压0.02~0.03MPa,设置搅拌速度100rpm/min,开始发酵。培养温度37±0.2℃,使用氨水作维持pH恒定(5.50-6.00)±0.02进行培养。
将发酵至终点的发酵液高速离心去除菌体,收集到的无细胞提取液经脱盐、浓缩和喷雾干燥制备得到粉剂。
所得粉剂再以10倍稀释复溶得到复溶液(相当于约10%干物质),将复溶液调pH值至7.0,通过测量抑菌圈直径大小判断其抑菌能力,特别是对荧光假单胞菌、铜绿假单胞菌、蜡样芽孢杆菌、单增李斯特菌、变异链球菌的抑菌能力。
此外,将副干酪乳酪杆菌PC646的无细胞提取液经脱盐、浓缩和喷雾干燥制备所得的粉剂以10倍(相当于约10%干物质)、20倍(相当于约5%干物质)、40倍(相当于约2.5%干物质)稀释复溶,分别将复溶液调pH值至7.0,通过测量抑菌圈直径大小,判断其对金黄色葡萄球菌的抑菌能力。
其中,将抑菌圈直径<8mm标记为“+”,直径为8~16mm标记为“++”,直径>16mm标记为“+++”,无抑菌活性的标记为“-”。
结果如表1所示:
表1副干酪乳酪杆菌PC646无细胞浓缩液抑菌谱
注:-表示无抑菌活性;+表示抑菌圈直径为<8mm;++表示抑菌圈直径为8~16mm;+++表示抑菌圈直径为>16mm
根据实验结果可知,副干酪乳酪杆菌PC646共对9种菌有抑菌作用。包括在革兰氏阴性菌中,对肠沙门氏菌(Salmonella enterica)、荧光假单胞菌(Pseudomonasfluorescens)、铜绿假单胞菌(Pseudomonas aeruginosa)、伴放线放线杆菌(Actinobacillus actinomycetemcomitans)、哈维氏弧菌(Vibrio harveyi)5种细菌有抑菌作用;在革兰氏阳性菌中,对金黄色葡萄球菌(Staphylococcus aureus)、单增李斯特氏菌(Listeria monocytogenes)、蜡样芽孢杆菌(Bacillus cereus)、变异链球菌(Streptococcus mutans)4种细菌有抑菌作用。
副干酪乳酪杆菌PC646对3种真菌均无抑菌作用。
其中,副干酪乳酪杆菌PC646的代谢产物喷干粉制剂10倍稀释复溶液对荧光假单胞菌、蜡样芽孢杆菌、单增李斯特菌、金黄色葡萄球菌均有抑菌作用,且对铜绿假单胞菌、变异链球菌有延缓生长作用(活性较弱),其抑菌圈直径如表2所示。
表2副干酪乳酪杆菌PC646代谢产物制剂抑菌活性测试
此外,副干酪乳酪杆菌PC646的代谢产物喷干粉制剂的10倍稀释复溶液、20倍稀释复溶液、40倍稀释复溶液,均对金黄色葡萄球菌有显著抑制作用,其结果如图1所示。
实施例3不同蛋白酶处理对副干酪乳酪杆菌PC646代谢产物制剂的抑菌活性影响
以金黄色葡萄球菌为指示菌,分别用蛋白酶K、胰蛋白酶、木瓜蛋白酶、蛋白酶E、胃蛋白酶对实施例2的粉剂的复溶液(20倍稀释,相当于约5%干物质)进行酶解(调复溶液至不同酶的最适作用pH值,其中,胃蛋白酶在酸性条件下,其余酶在中性条件下,使每种酶的终浓度为1mg/mL,以蒸馏水代替酶液作为对照组,37℃孵育2h后将pH值调回7.0,过0.22μm滤器过滤除菌),依照实施例2方法测试不同蛋白酶处理对抑菌活性的影响。
结果显示,副干酪乳酪杆菌PC646代谢产物制剂的复溶液经木瓜蛋白酶酶解后,对金黄色葡萄球菌的抑菌活性完全消失;经其它四种蛋白酶处理后,抑菌活性没有明显变化,结果如表3、图2所示。
表3不同蛋白酶处理对副干酪乳酪杆菌PC646代谢产物制剂的抑菌活性影响
综上,副干酪乳酪杆菌PC646代谢产物制剂的抑菌活性物质可能为蛋白类成分,其对木瓜蛋白酶较敏感,但其他蛋白酶并未显著影响其抑菌活性。
实施例4副干酪乳酪杆菌PC646代谢产物制剂在熟牛肉防腐中的应用
从当地超市购买的新鲜牛肉切成小块,每块重量约1.5克,121℃灭菌15分钟,获得无菌熟牛肉样品,每块重量约1g。
将熟牛肉样品在PC646代谢产物制剂溶液(10%浓度)中浸泡30秒,记为实验组,对照组不做处理。将对照组和实验组样品接种金黄色葡萄球菌,接种量约102CFU/g,接种后密封保存放置于4℃冰箱,分别在0、3、6和9天取样进行微生物分析、酸碱度测定、挥发性盐基氮(TVB-N)和硫代巴比妥酸反应物(TBARS)测定。
结果显示,与对照组相比,经副干酪乳酪杆菌PC646代谢产物制剂处理后的实验组金黄色葡萄球菌数量在贮藏期间没有显著增加,而对照组的菌落数量随时间显著增加,如图3所示。
与对照组相比,经副干酪乳酪杆菌PC646代谢产物制剂处理后的熟牛肉的pH值没有显著差异,且在储存时间里,实验组和对照组的pH均没有显著性变化,如图4所示。
与对照组相比,经副干酪乳酪杆菌PC646代谢产物制剂处理使牛肉颜色变暗,但在储存的时间里总色差没有显著变化,而对照组在储存期间总色差有显著性变化。结果说明代谢产物制剂在肉牛肉的贮藏期间具有肉色稳定效果,如图5所示。
与对照组相比,经副干酪乳酪杆菌PC646代谢产物制剂处理的实验组的挥发性盐基氮含量显著降低,说明副干酪乳酪杆菌PC646代谢产物制剂在储存期间能够显著延缓熟牛肉中挥发性盐基氮的发展,如图6所示。
与对照组相比,副干酪乳酪杆菌PC646代谢产物制剂加入可以显着降低熟牛肉的硫代巴比妥酸反应物的值,如图7所示,表明副干酪乳酪杆菌PC646代谢产物抑制了熟牛肉在储存期间脂质的过氧化。
以上结果表明本发明的副干酪乳酪杆菌PC646具有抑菌潜力,其发酵后的无细胞提取液经脱盐、浓缩和喷雾干燥制备所得的抑菌制剂对于常见致病菌,特别是金黄色葡萄球菌,具有显著的抑制效果且可以用于食品防腐保鲜。本发明为该菌株及其抑菌活性制剂的深入研究和开发利用提供了一种全新的思路。
Claims (4)
1.副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646在抑制致病菌中的应用,所述致病菌是肠沙门氏菌(Salmonella enterica)、荧光假单胞菌(Pseudomonasfluorescens)、铜绿假单胞菌(Pseudomonas aeruginosa)、伴放线放线杆菌(Actinobacillus actinomycetemcomitans)或哈维氏弧菌(Vibrio harveyi);在革兰氏阳性菌中,对金黄色葡萄球菌(Staphylococcus aureus)、单增李斯特氏菌(Listeriamonocytogenes)、蜡样芽孢杆菌(Bacillus cereus)或变异链球菌(Streptococcusmutans),所述副干酪乳酪杆菌PC646保藏于中国微生物菌种保藏管理委员会普通微生物中心,其保藏编号为CGMCC No.23515。
2.副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646在食品添加剂中的应用,所述副干酪乳酪杆菌PC646保藏于中国微生物菌种保藏管理委员会普通微生物中心,其保藏编号为CGMCC No.23515。
3.根据权利要求2所述的应用,其特征在于所述食品添加剂是食品防腐剂。
4.一种具有抑菌特性的益生菌粉剂的制备方法,所述方法包括以下步骤:
(1)种子培养
将冷冻保存在-80℃的副干酪乳酪杆菌(Lacticaseibacillus paracasei)PC646接种至MRS液体培养基中,于37℃培养24h,以体积分数2%的接种量连续活化至第3代,培养18h后,得到种子液;
(2)扩大培养
将所述种子液以体积比5%的接种量接入强化MRS液体培养基中,然后加入氨水或氢氧化钠调整发酵液初始pH至6.50±0.02,空气保压0.02~0.03MPa,设置搅拌速度100rpm/min,再温度37±0.2℃条件下进行发酵,使用氨水作维持pH恒定(5.50-6.00)±0.02;
(3)发酵液的后处理
将步骤(2)发酵至终点的发酵液高速离心去除菌体,收集到的无细胞提取液经脱盐、浓缩和喷雾干燥制备得到粉剂。
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CN116814481B (zh) * | 2023-06-09 | 2024-04-09 | 内蒙古农业大学 | 一株源自酸马奶的益生副干酪乳酪杆菌pc646及其人工智能筛选方法 |
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