Open L-lactic acid fermentation of food refuse using thermophilic Bacillus coagulans and fluorescence in situ hybridization analysis of microflora

In the production of commercially useful poly-L-lactic acid plastic from biomass wastes, a feasible fermentation process to produce optically active L-lactic acid would be required. Here, model kitchen refuse (MKR) was inoculated with Bacillus coagulans NBRC12583 under nonsterilized openculture conditions. At temperatures below 45 degrees C, a racemic mixture of D- and L-lactic acids was accumulated, whereas only L-lactic acid was selectively accumulated by incubation at 50-65 degrees C. At 45 degrees C, the results of fermentation could not be consistently reproduced. To analyze microflora in this type of mixed culture system, whole-cell fluorescence in situ hybridization (FISH) using 16S rRNA-targeted oligonucleotide probes for B. coagulans, Bcoa191, and LAC722(L), a group-specific probe for a wide range of mesophilic lactic acid bacteria was applied. The dominancy of mesophilic lactic acid bacteria at lower temperatures, and that of B. coagulans at higher temperatures were confirmed. By using a saccharified liquid of collected kitchen refuse, 86 g/l of L-lactic acid was accumulated under nonsterile conditions by a 5-d incubation at 55 degrees C, pH 6.5, with 53% carbon yield and 97% optical purity. To conclude, high temperature open lactic acid fermentation is a simple and promising method for producing high-grade L-lactic acid from biomass waste, and FISH analysis of such mixed-culture systems is helpful for monitoring the microflora in these cultures.     

Racemization of l-lactic acid in pH-swing open fermentation of kitchen refuse by selective proliferation of Lactobacillus plantarum

We have shown that stable lactic acid fermentation of model kitchen refuse occurs with intermittent pH adjustment under nonsterilized conditions. Nonetheless, the optical activity of the accumulated lactic acid was low, which is disadvantageous for the production of high-quality poly-l-lactic acid. Here, we attempt to increase optical purity by introducing l-lactic acid-producing strains under nonsterilized conditions and demonstrate that the inoculation of Lactobacillus rhamnosus or Lactococcus lactis, both of which are l-lactic acid producers, is partially effective in the early fermentation stage, but does not improve the final optical purity of the accumulated lactic acid. We confirmed by fluorescence in situ hybridization using group-specific and species-specific 16S rDNA probes that this is due to the selective proliferation of naturally existing L. plantarum. L. plantarum KY-1, which is isolated from model kitchen refuse, showing lactic acid racemase activity, as well as d-lactate dehydrogenase activity, in its membrane fraction. We conclude that racemase activity associated with L. plantarum is the main cause of decreased optical purity in the accumulated lactic acid.     

原位产物分离技术在乳酸发酵中的应用

介绍了乳酸发酵中常用的几种原位产物分离技术——电渗析发酵、膜法发酵、提取发酵、吸附发酵。原位产物分离技术克服了传统发酵过程中发酵液里过高的乳酸盐对乳酸菌的抑制作用。     

原位分离技术在乳酸发酵中的应用

论述了乳酸发酵中几种常用的原位分离技术--膜发酵法、电渗析发酵、萃取发酵、吸附发酵,介绍了各自的应用特点以及研究现状.     

Thermotolerant Bacillus licheniformis TY7 produces optically active l-lactic acid from kitchen refuse under open condition

A thermotolerant l-lactic-acid-producing bacterium was isolated and identified as Bacillus licheniformis TY7. TY7 shows optimum growth at pH 6.5 at 30 degrees C and normal growth up to 65 degrees C. Using nonsterile kitchen refuse at 50 degrees C, the strain produced 40 g/ll-lactic acid with 97% optical activity and 2.5 g/lxh productivity.     

Hydrogen production by fermentation using acetic acid and lactic acid

Microbial hydrogen production from sho-chu post-distillation slurry solution (slurry solution) containing large amounts of organic acids was investigated. The highest hydrogen producer, Clostridium diolis JPCC H-3, was isolated from natural environment and produced hydrogen at 6.03+/-0.15 ml from 5 ml slurry solution in 30 h. Interestingly, the concentration of acetic acid and lactic acid in the slurry solution decreased during hydrogen production. The substrates for hydrogen production by C. diolis JPCC H-3, in particular organic acids, were investigated in an artificial medium. No hydrogen was produced from acetic acid, propionic acid, succinic acid, or citric acid on their own. Hydrogen and butyric acid were produced from a mixture of acetic acid and lactic acid, showing that C. diolis. JPCC H-3 could produce hydrogen from acetic acid and lactic acid. Furthermore, calculation of the Gibbs free energy strongly suggests that this reaction would proceed. In this paper, we describe for the first time microbial hydrogen production from acetic acid and lactic acid by fermentation.     

乳酸发酵海藻酸钠果冻的生产工艺

介绍乳酸发酵海藻酸钠果冻的生产原理、配方及工艺。     

直接乳酸发酵工艺研究进展

对以淀粉质为原料进行直接乳酸发酵的国内外研究现状进行了综述.介绍了利用米根霉等真菌和解淀粉乳酸细菌进行直接乳酸发酵.以及通过添加降解酶进行同步糖化发酵的三种直接乳酸发酵工艺.指出通过筛选和构建更加高效的具有解淀粉活性的乳酸细菌和真菌,同时通过获得耐受更高温度的乳酸菌来提高同步糖化发酵的温度,进而使直接乳酸发酵工艺更加高效.     

Rapid and sensitive in situ hybridization method for detecting and identifying lactic acid bacteria in wine

A rapid and sensitivein situhybridization method, using total genomic DNA probes, can specifically detect and identify the lactic acid bacteria in wine. The total genomic DNA probes were labelled with digoxygenin (DIG) by random priming and hybridized with the genomic DNA of the bacteria to be identified. The hybrids were detected with fluorescent anti-DIG Fab-fragments or with enzyme-conjugated anti-DIG Fab-fragments. The cell wall of the Gram-positive lactic acid bacteria was made permeable with a lysozyme treatment to allow the accessibility of the probe to the target DNA and the penetration of anti-DIG antibodies which specifically bound to their antigens. The total genomic DNA probes exhibited the same specificity with dot-blot andin situhybridization. Thein situhybridization technique made possible the control of the bacterial population in wine at different stages of vinification of storage, and the identification of lactic acid bacteria which can cause wine spoilage.     

乳酸啤酒发酵的研究

从啤酒厂的优质酵母泥中分离筛选出一株发酵性能良好的卡尔酵母HF1。以HF1为出发菌株,植物乳杆菌培养液作为生物酸化法糖化麦汁的pH值调节剂,在微型啤酒发酵罐中对生物酸化麦汁进行发酵,发酵后的嫩啤酒达到了GB4927-2001优质啤酒的标准。     

Molecular characterization of lactic acid bacteria and in situ amylase expression during traditional fermentation of cereal foods

Lactic acid bacteria play an important role in traditional fermented foods consumed in different countries. Study of their taxonomic structure and diversity is necessary for starter culture selection, improved safety and nutritional enhancement. To achieve these objectives, microbial genomic typing methods were used to study genetic differences of autochthonous bacteria and their distribution in two traditional African fermented cereal foods. A total of 85 predominant bacterial species were isolated from ogi and kunu-zaki obtained from Northern and Southern geographical region of Nigeria. They were identified using combination of 16S rRNA gene sequencing, multilocus sequence analysis (MLSA) based on rpoA, pheS and atpA genes as well as M13-PCR gel fingerprints. The results showed that Lactobacillus fermentum was the most frequently isolated species in ogi (71.4%) and kunu-zaki (84.5%). Other species of lactic acid bacteria (LAB) identified were Lactobacillus plantarum, Streptococcus gallolyticus subsp. macedonicus and Pediococcus pentosaceus. Non lactic acid bacteria isolated from these foods were species belonging to the Bacillus and Staphylococcus. Non-metric multidimensional scaling (nMDS) analysis of the M13-PCR fingerprints for LAB strains showed clonal diversity among strains of the same species. In vitro and in situ expression of amylase gene during fermentation by amylolytic L. plantarum ULAG11 was detected, indicating the potential usefulness of such species for development of starter cultures and for controlled fermentation processes.     

Ultrafiltration flux of acid whey obtained by lactic acid fermentation

Acid whey was prepared from reconstituted ultra-low heat skimmed milk powder fermented with Lactobacillus helveticus, clarified by centrifugation, pH adjusted and ultrafiltered (cut-off limit 10 or 30 kDa) at 50 °C to a volume reduction ratio of 10. Centrifugation primarily removed the serum albumin and, to a lesser extent, immunoglobulins, considered the most critical membrane foulants during ultrafiltration of acid whey. Below pH 3.9 the flux improvement was directly dependent on pH lowering. At pH 3.9 a deviation point in the relationship between pH and flux of acid whey was observed. Above this pH a severe flux decline was detected and at pH 4.6 the fermented acid whey was non-filterable. Largest flux increase was obtained at a transmembrane pressure of 2.0 bar and 50 °C. Under these conditions the permeation flux achieved using a 30 kDa membrane was about 34 ± 5 kg h⁻¹m⁻², comparable with that for sweet whey.     

Pressure-resistant acclimation of lactic acid bacteria from a natural fermentation product using high pressure

Twenty colonies of Pediococcus acidilactici, Lactobacillus brevis, Lactiplantibacillus plantarum and Latilactobacillus curvatus were isolated from fermentation broth of pickled pepper by using MRS agar. The strains were acclimated using high pressure from 300 to 400 MPa for 1 to 5 min. For one generation of acclimation, the survived cells after first high pressure treatment were then activated by culturing on MRS agar for two days for the next high pressure treatment. After acclimation for 30 generations, the survival ratio of L. plantarum and L. curvatus was increased by 138 and 1222 times at 400 MPa for 1 and 3 min, respectively. Weibull model could appropriately fit the survival curves of L. plantarum and L. curvatus treated by high pressure. The time to the first decimal reduction of these two strains after acclimation was 3.5 and 1.8 times, respectively, longer than before acclimation at 300 MPa. From electron microscopy photographs, the acclimated L. curvatus was more curved, smaller and its cell wall was thicker than the unacclimated L. curvatus. After HPP treatment, L. plantarum and L. curvatus have higher cell integrity, smoother cell surface, more uniform protoplasm and smaller cavities in the cell protoplasm compared with unacclimated strains, suggesting that high pressure acclimation introduced some modification to bacteria intrinsic structures and increased their pressure resistance. Moreover, the fermentation performance including glucose utilization capacity and lactic acid production of the acclimated L. curvatus was respectively improved by 9.6% and 9.4% at 37 °C for 24 h in MRS medium.     

浓香型白酒发酵中乳酸含量动态变化与相关微生物研究

在浓香型白酒固体发酵中,乳酸含量变化趋势具有一定的规律性.从不同阶段分离的微生物中,鉴别、统计产乳酸以及利用乳酸牛长的微牛物种类和数量.结果表明,发酵前1周内,乳酸利用菌的数量和性能略占优势(达到108水平),使乳酸含量保持较低水平(2%～2.5%),1周以后,乳酸利用菌数量下降明显(106水平),与产乳酸菌数量相近,使乳酸含量维持在较高水平.研究结果表明,既能产乳酸,又能利用乳酸的菌株,对固态发酵的影响结果与其产乳酸能力呈正相关.     

不同乳酸菌在毛酸浆发酵中的特性研究

为比较嗜热链球菌（Lactobacillus plantarum）（L1）、保加利亚乳杆菌（Lactobacillus bulgaricus）（L2）、瑞士乳杆菌（Lactobacillus helveticus）（L3）、植物乳杆菌（Lactobacillus plantarum）（L4）、短乳杆菌（Lactobacillus brevis）（L5）和干酪乳杆菌（Lacbobacillus casei）（L6）在毛酸浆发酵中的特性,研究发酵过程中总酸含量、活菌数和色泽的变化。结果表明,发酵过程中,6种乳酸菌活菌数均能维持在8.0 lg（CFU/mL）以上,生长良好,其中乳酸菌L4、L5和L6在毛酸浆中产酸能力较好,最高总酸含量分别为11.85 g/L、10.95 g/L、10.45 g/L,并均能很好地保持发酵液本身的颜色,色差值均＜54.51;当乳酸菌L4、L5和L6按不同接种比例复合发酵毛酸浆果汁时,各处理间活菌数和发酵液颜色变化基本一致,当乳酸菌L4∶L6为3∶2（V/V）进行复合发酵时,最终所得毛酸浆发酵液的总酸含量（17.91 g/L）最高。     

Exploitation of vegetables and fruits through lactic acid fermentation

Lactic acid fermentation represents the easiest and the most suitable way for increasing the daily consumption of fresh-like vegetables and fruits. Literature data are accumulating, and this review aims at describing the main features of the lactic acid bacteria to be used for fermentation. Lactic acid bacteria are a small part of the autochthonous microbiota of vegetables and fruits. The diversity of the microbiota markedly depends on the intrinsic and extrinsic parameters of the plant matrix. Notwithstanding the reliable value of the spontaneous fermentation to stabilize and preserve raw vegetables and fruits, a number of factors are in favour of using selected starters. Two main options may be pursued for the controlled lactic acid fermentation of vegetables and fruits: the use of commercial/allochthonous and the use of autochthonous starters. Several evidences were described in favour of the use of selected autochthonous starters, which are tailored for the specific plant matrix. Pro-technological, sensory and nutritional criteria for selecting starters were reported as well as several functional properties, which were recently ascribed to autochthonous lactic acid bacteria. The main features of the protocols used for the manufacture of traditional, emerging and innovative fermented vegetables and fruits were reviewed. Tailored lactic acid bacteria starters completely exploit the potential of vegetables and fruits, which enhances the hygiene, sensory, nutritional and shelf life properties.     

乳酸菌发酵柿汁的制备工艺

介绍了以柿子汁为原料制备乳酸菌发酵饮料的工艺流程,并对原材料和发酵产品的一些营养指标进行了测量。实验表明,乳酸菌发酵的最佳条件是发酵温度30℃、接种量6%、菌种AS1.1482∶6038=2∶1。产品既保留了部分柿子汁的原有风味,又具有乳酸发酵形成的特殊风味,营养价值较高,有一定的保健作用。     

乳酸菌发酵黑豆乳理化性质的研究

黑豆经浸泡、打浆、杀菌等处理后 ,制成黑豆乳 ,接种乳酸菌进行发酵实验。结果表明 :经乳酸菌发酵后的黑豆乳 ,除具有一般乳酸产品所共有的适口酸味外 ,还具有黑豆特有的香气。但产物酸度偏低。