高密度发酵和真空冷冻干燥工艺对乳酸菌抗冷冻性的影响

经真空冷冻干燥得到的乳酸菌发酵剂存活率和后期的低温贮藏稳定性与诸多因素相关。本文综述了制备乳酸菌发酵剂过程中高密度发酵和真空冷冻干燥工艺的不同对乳酸菌抗冷冻性的影响,其中高密度发酵过程中的培养基组分、培养温度、发酵恒定pH、中和剂的选择、菌体收获时期和发酵结束后处理以及真空冷冻干燥过程中保护剂的添加、预冷冻处理等是影响乳酸菌抗冷冻性的重要因素。通过对这些相关因素的综述分析,为提高乳酸菌发酵剂的冻干存活率和后期的低温贮藏稳定性提供新的思路,且应用抗冷冻性强、活力高的乳酸菌发酵剂对有效提高乳制品的质量和企业的经济效益意义重大。     

菌种冷冻干燥保藏的影响因素

菌种资源保藏是微生物学及相关学科研究的基础.冷冻干燥保藏法是菌种保藏最有效的方法之一,为进一步提高菌种保藏质量人们进行了大量的研究.本文介绍了菌种冷冻干燥保藏方法的原理和优点,同时详细介绍了菌种冷冻干燥保藏方法的影响因素.     

活菌制剂冻干保护剂的研究进展

冷冻干燥技术广泛应用于细菌性活疫苗生产和菌种保藏等方面。细菌细胞在冷冻干燥过程中会出现损伤,甚至死亡。通过添加适合的冻干保护剂可以最大程度减小细胞损伤,保持活菌制剂的活力和性能。就冻干过程对细菌的损伤机制、冻干保护剂的作用机理以及保护剂的筛选方法等方面进行了阐述,对活菌制剂保护剂的筛选有所启示。     

乳酸菌的抗冷冻性及耐受机理

乳酸菌发酵剂在工业生产过程中,会受到冷冻的刺激,如真空冷冻干燥及后期的低温保藏,此外,发酵乳制品的保藏和干酪的成熟过程也都在低温中进行。这些均会对乳酸菌发酵剂及发酵乳制品质量产生一定的影响。因此,掌握乳酸菌在冷冻条件下的反应机理有助于优化发酵剂和发酵乳制品在工业生产中的冷冻、发酵和贮藏条件,从而提高产品质量和生产效益。本文对乳酸菌的抗冷冻性及机理进行了分析,并对发酵剂的保护提出具体措施。     

菌种冷冻干燥保藏的影响因素

菌种资源保藏是微生物学及相关学科研究的基础。冷冻干燥保藏法是菌种保藏最有效的方法之一, 为进一步提高菌种保藏质量人们进行了大量的研究。本文介绍了菌种冷冻干燥保藏方法的原理和优点, 同时详细介绍了菌种冷冻干燥保藏方法的影响因素。     

冷冻干燥对乳酸菌细胞膜通透性的影响

对细胞膜通透性变化的研究是认识冷冻干燥过程对乳酸菌损伤机理的途径之一。用荧光探针检测冻干过程前后细胞内H+和Ca2+浓度的变化, 可以精确的表征细胞膜通透性的改变。利用荧光探针BCECF-AM和Fluo3-AM对德氏乳杆菌保加利亚亚种在冻干前后的细胞膜通透性进行研究, 并对比菌种在冻干过程中的活力损失, 发现细胞膜在冻干前后通透性有显著增加, 并与活力的损失成反相关关系。说明在冻干过程中细胞受到了生理性损伤, 细胞膜通透性的改变可能是导致乳酸菌在冻干过程中致死和失活的原因之一。     

降胆固醇益生菌发酵剂的制备

以前期筛选出来的降胆固醇益生菌嗜酸乳杆菌S-59为出发菌株，对冻干发酵剂的制备工艺进行研究和优化。结果表明：菌悬液的离心收集条件为5 000 r/min、20 min；通过单因素试验和响应面优化试验，确定冻干保护剂为脱脂乳8.95%、海藻糖5.24%、甘油4.54%，细胞存活率可达90.1%。冻干发酵剂的保藏条件为在真空条件下-4℃保藏。     

抗绿脓杆菌鸡卵黄色免疫蛋白的冷冻干燥研究

介绍了冷冻干燥技术的原理、抗绿脓杆菌(Pseudomonas aeruginosa,简称PA)鸡卵黄免疫球蛋白(Immunoglobulin of Yolk,IgY)的冷冻干燥工艺过程及其参数.通过实验,获得了抗-PA IgY的冻干曲线;经间接血球凝集实验检测,抗-PAIgY的冻干品的活性为1128;在4℃和25℃下,抗-PAIgY的冻干品保存6个月,其活性不变.     

微生物冷冻干燥的抗性机理

微生物冷冻干燥的抗性机理陈声明,吕琴（浙江农业大学环保系,杭州３１００２９）微生物冷冻干燥法是长期保藏菌种的较好方法之一。因其便于批量生产、长期保藏成活力高,保藏和运输设备简单而成为一种普遍使用的保藏方法,已广泛应用于保存大部分细菌、真菌和一些病毒。...     

不同培养基对酒酒球菌SD-2a存活率及膜脂肪酸组分的影响

[目的] 为获得高效的葡萄酒乳酸菌发酵剂,本文研究了3种具有不同pH缓冲能力的培养基对酒酒球菌接种存活率、冻干存活率及细胞膜脂肪酸组分的影响.[方法]采用平板计数法测定菌体的接种存活率、冻干存活率;并采用GC/MS色谱方法测定收获菌体细胞膜脂肪酸组分.[结果]实验结果表明,没有添加苹果酸的ATB培养基,其pH缓冲能力弱.分别与FMATB和MATB培养基相比,ATB培养基培养获得的菌体,其接种模拟酒培养基后的存活率提高了20.3%和40.2%,其冷冻干燥存活率提高了48.5%和68.3%,其细胞膜中C19cyc11的相对含量提高了10.0%和36.8%,其细胞膜U/S值提高了20.4%和45.2%.[结论]本文推测ATB培养基培养所得菌体,由于自我酸胁迫反应,增强了其对葡萄酒胁迫因素及冷冻干燥的抗性,而该反应与菌体细胞膜脂肪酸组分的变化密切相关.故ATB培养基更适合于酒酒球菌SD-2a发酵剂的制备.     

Survival of Escherichia coli during drying and storage in the presence of compatible solutes

Five different compatible solutes, sucrose, trehalose, hydroxyectoine, ectoine, and glycine betaine, were investigated for their protective effect on Escherichia coli K12 and E. coli NISSLE 1917 during drying and subsequent storage. Two different drying techniques, freeze-drying and air-drying, were compared. The highest survival rate was observed when the non-reducing disaccharides sucrose (for E. coli K12) and trehalose (for E. coli NISSLE 1917) were added. The two tetrahydropyrimidines, hydroxyectoine and ectoine, gave protection to freeze-dried E. coli NISSLE 1917 whereas E. coli K12 was protected only by hydroxyectoine. Glycine betaine seemed to be harmful for both strains of E. coli with both drying techniques. Air0drying gave much better survival rates than freeze-drying. The two strains of E. coli differed in their ability to take up compatible solutes.     

Survival curves for microbial species stored by freeze-drying

The survival of a variety of species of microorganism following storage for up to 20 years has been analyzed. The organisms were freeze-dried, sealed in ampoules under vacuum (<1 Pa) and stored in the dark at 5 degrees C. The yeast that was tested, Saccharomyces cerevisiae, showed only 8% survival when recovered shortly after freeze-drying, but subsequent loss during storage was the least among all the tested microorganisms. The decrease in the logarithm of survival per year (log survival) was -0.010, which corresponds to a survival rate of 97.7% per year. The Gram-negative bacteria tested, Escherichia coli, Pseudomonas putida, and Enterobacter cloacae, showed 42.6, 33.5, and 50.8% survival shortly after freeze-drying, which was higher than the corresponding survival of S. cerevisiae, but the subsequent loss during storage was greater than S. cerevisiae, the log survival figures being -0.041, -0.058, and -0.073 per year. These values correspond to survival rates of 91.0, 87.5, and 84.5% each year. The Gram-positive bacteria tested, Lactobacillus acidophilus and Enteroccoccus faecium, showed 62.5 and 85.2% survival shortly after freeze-drying, which was even higher than that of the Gram-negative species, and these organisms also showed better survival during storage than Gram-negative bacteria; their log survival rates were -0.018 and -0.016 per year, which corresponded to survival rates of almost 96% per year. Comparison of these results with other published data for different drying conditions suggests that survival during storage is strongly influenced by the degree of vacuum under which the ampoules were sealed. The excellent survival after freeze-drying of each species might be attributable to the high level of desiccation and to sealing under vacuum.     

Effects of various sugars added to growth and drying media upon thermotolerance and survival throughout storage of freeze-dried Lactobacillus delbrueckii ssp. bulgaricus

The aim of this research effort was to investigate the role of various sugar substrates in the growth medium upon thermotolerance and upon survival during storage after freeze-drying of Lactobacillus bulgaricus. Addition of the sugars tested to the growth medium, and of these and sorbitol to the drying medium (skim milk) was investigated so as to determine whether a relationship exists between growth and drying media, in terms of protection of freeze-dried cells throughout storage. The lowest decrease in viability of L. bulgaricus cells after freeze-drying was obtained when that organism was grown in the presence of mannose. However, L. bulgaricus clearly survived better during storage when cells had been grown in the presence of fructose, lactose or mannose rather than glucose (the standard sugar in the growth medium). A similar effect could not be observed in terms of thermotolerance; in this case, the growth medium supplemented with lactose was found to yield cells bearing the highest heat resistance. Supplementation of the drying medium with glucose, fructose, lactose, mannose or sorbitol led in most cases to enhancement of protection during storage, to a degree that was growth medium-dependent.     

Influence of microencapsulation and spray drying on the viability of Lactobacillus and Bifidobacterium strains

Improved production methods of starter cultures, which constitute the most important element of probiotic preparations, were investigated. The aim of the presented research was to analyse changes in the viability of Lactobacillus. acidophilus and Bifidobacterium bifidum after stabilization (spray drying, liophilization, fluidization drying) and storage in refrigerated conditions for 4 months. The highest numbers of live cells, up to the fourth month of storage in refrigerated conditions, of the order of 10(7) cfu/g preparation were recorded for the B. bifidum DSM 20239 bacteria in which the N-Tack starch for spray drying was applied. Fluidization drying of encapsulated bacteria allowed obtaining a preparation of the comparable number of live bacterial cells up to the fourth month of storage with those encapsulated bacteria, which were subjected to freeze-drying but the former process was much shorter. The highest survivability of the encapsulated L. acidophilus DSM 20079 and B. bifidum DSM 20239 cells subjected to freeze-drying was obtained using skimmed milk as the cryoprotective substance. Stabilization of bacteria by microencapsulation can give a product easy to store and apply to produce dried food composition.     

Optimisation and comparison of liquid and dry formulations of the biocontrol yeast <Emphasis Type="Italic">Pichia anomala</Emphasis> J121

The biocontrol yeast Pichia anomala J121 can effectively reduce mould growth on moist cereal grains during airtight storage. Practical use of microorganisms requires formulated products that meet a number of criteria. In this study we compared different formulations of P. anomala. The best way to formulate P. anomala was freeze-drying. The initial viability was as high as 80%, with trehalose previously added to the yeast. Freeze-dried products could be stored at temperatures as high as 30 °C for a year, with only a minor decrease in viability. Vacuum-drying also resulted in products with high storage potential, but the products were not as easily rehydrated as freeze-dried samples. Upon desiccating the cells using fluidised-bed drying or as liquid formulations, a storage temperature of 10 °C was required to maintain viability. Dependent on the type of formulation, harvesting of cells at different nutritional stresses affected the initial viabilities, e.g. the initial viability for fluidised-bed-dried cells was higher when the culture was fed with excess glucose, but for freeze-drying it was superior when cells were harvested after depletion of carbon. Using micro-silos we found that the biocontrol activity remained intact after drying, storage and rehydration for all formulations.     

Survival kinetics of lactic acid starter cultures during and after freeze drying

Survival kinetics of lactic acid starter cultures were modeled considering the microorganism and external medium interfacial area as the critical factors determining the resistance of the microorganisms to freeze-drying. Surviving fraction of the microorganisms increased with the increasing biomass concentration during freeze-drying, and this is attributed to the mutual shielding effect of the microorganisms against the severe conditions of the external medium. Survival of the microorganisms over the storage period after freeze drying was enhanced by the presence of dead microorganisms which reduce the interfacial area between the live cells and the external medium. Streptococcus thermophilus was found to be more resistant to freeze-drying conditions than Lactobacillus bulgaricus. Storage under vacuum or nitrogen was superior to storage under air. Poor survival rates under air was attributed to oxygen diffusion into the dry cells through the interfacial area.     

Effect of drying on conidial viability of Penicillium frequentans, a biological control agent against peach brown rot disease caused by Moniliniaspp

The effects of drying methods (freeze-, spray-, and fluid bed-drying) on viability of Penicillium frequentans conidia were compared. Viability, estimated by germination of fluid bed- and freeze-dried conidia, was similar to that of fresh conidia. Skimmed milk alone, or in combination with other protectants, was added to conidia before freeze-drying. After the freeze-drying process, all protectants used, except glycerol improved conidial viability. Freeze-dried P. frequentans conidia did not maintain viability after 30 days of storage at room temperature, while conidia dried by fluid bed-drying showed 28% viability following 180 days after drying. This work also demonstrated a relationship between conidial viability after 1 year of storage at room temperature, moisture content after fluid bed-drying and initial weight of sample. Conidial moisture contents must be reduced to 5-15% for optimal storage at room temperature. P. frequentans conidia dried by fluid bed-drying were as effective as fresh conidia in controlling brown rot of peaches.     

Factors affecting the viability and properties of microorganisms in different preservation methods

The review of literature devoted to the influence of the different methods of longterm preservation on the survival, physiological and biochemical properties of microorganisms: at low and ultralow temperatures, freeze-drying, drying, storage under the mineral oil etc. is given. The microorganisms viability depends on their nature, age and density of population, storage and recovery conditions of cells. Some features of the industrial microorganisms storage have been marked. The different hypotheses concerning the mechanism of preservation, injury and reactivation of microorganisms under the action of external factors during their storage are being discussed.     

益生菌冷冻干燥高活性保护机制研究进展

摄取足量益生菌有助于维持肠道微生物群落的稳态,对维持人体肠道健康具有重要意义。然而,在工业化应用中,益生菌抗逆能力较弱且对储存条件要求高,导致益生菌产品对运输和活性维持条件要求较高,这些产业需求对高活力益生菌的制备工艺提出了挑战。干燥处理常应用于保持益生菌活性和稳定性,其中冷冻干燥技术应用最广泛,但冻干过程中益生菌会受到各类环境压力的刺激,引起细胞损伤甚至死亡。因此,可以显著提高益生菌存活率的冻干保护剂成为目前益生菌工业应用的研究热点。本文从益生菌常用及新发现的冻干保护剂种类及其作用机制进行了系统归纳,对菌株冻干后细胞存活率的影响因素进行全面综述,并对冻干保护剂研究方向进行了展望,旨在为高活力益生菌冻干菌粉的研制提供理论支持。     

Survival rate of microbes after freeze-drying and long-term storage

The survival rates of 10 species of microorganisms were investigated after freeze-drying and preserving in a vacuum at 5 degrees C. The survival rates varied with species. The survival rates immediately after freeze-drying were different among yeast, gram-positive bacteria, and gram-negative bacteria, and the change in the 10-year survival rate was species-specific. The survival rate of yeast, Saccharomyces cerevisiae, was about 10% immediately after drying, and the rate did not decrease significantly during the 10-year storage period. Survival rates after the drying of gram-positive bacteria, i.e., Brevibacterium flavum, B. lactofermentum, Corynebacterium acetoacidophilum, C. gultamicum, and Streptococcus mutans, were around 80%. The survival rate of Brevibacterium and Corynebacterium did not decrease greatly during the storage period, whereas the rate of S. mutans decreased to about 20% after 10 years. Survival rates after the drying of gram-negative bacteria, i.e., Escherichia coli, Pseudomonas putida, Serratia marcescens, and Alcaligenes faecalis, were around 50%. The survival rate decreased for the first 5 years and then stabilized to around 10% thereafter.