Production of canthaxanthin by extremely halophilic bacteria

Soil samples from a salt farm were used as a source for the isolation of carotenoid-producing bacteria. The conditions for optimum growth and carotenoid production were established for the isolated bacteria. Carotenoids were analysed by spectrophotometry and High Performance Liquid Chromatography (HPLC). Thirty-one red extremely halophilic bacteria were isolated from saline soil samples collected from a salt farm in Alexandria, Egypt. Among the isolated strains, strain TM exhibited the highest carotenoid-producing ability. Maximum growth of strain TM occurred in the presence of high concentrations of sodium chloride and magnesium sulfate. Growth did not occur when NaCl concentration was lower than 10% and the cells lysed at this concentration. Optimum growth of and carotenoid production by strain TM were realized at 37 degrees C in the presence of 1% yeast extract, 0.75% casamino acids, 25% NaCl, 4% MgSO4, 0.2% KCl and at pH 7.2 with shaking for 6 d. Strain TM produced 2.06 mg total carotenoids g(-1) dry cells, including 0.06 mg of beta-carotene and 0.70 mg of canthaxanthin. This is the first report of an extremely halophilic bacterium that produces canthaxanthin.     

Production of pyroglutamic acid by thermophilic lactic acid bacteria in hard-cooked mini-cheeses

Pyroglutamic acid is present in high amounts (0.5g/ 100g) in many cheese varieties-and particularly in extensively ripened Italian cheeses such as Grana Padano and Parmigiano Reggiano. An in vivo model system for cooked mini-cheese production and ripening acceleration was set up to demonstrate the ability of thermophilic lactic acid bacteria, used as a starter, to produce pyroglutamic acid (pGlu). In mini-cheeses stored at 38 and 30 degrees C for up to 45 d, all starters tested produced different amounts of pGlu. In descending order of pGlu production, the bacteria analyzed were: Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and Lactobacillus delbrueckii subsp. lactis. Evidence for the presence of glutamine to pGlu cyclase activity in lactic acid bacteria was provided. Cell lysates obtained from cultures of L. helveticus, L. delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis, and S. thermophilus showed the ability to cyclize glutamine to pGlu, resulting in processing yields from 1.4 to 30.3%, depending on the subspecies. Formation of pGlu from free glutamine appeared to be similar to that observed using a glutamine-glutamine dipeptide substrate. Under the experimental conditions applied, pGlu aminopeptidase activity was only detected in L. helveticus. Thus, pGlu formation in long-ripened cooked cheese may depend on the activity of thermophilic lactic acid bacteria.     

Nitrite production by thermophilic aerobic bacteria isolated during the manufacture of milk powder

Nitrite contamination in milk powder is hypothesised to be caused by thermophilic bacteria that form biofilms in milk powder manufacturing plants. Regulatory limits in some countries have made nitrite contamination in milk powder an issue influencing product sales. In this study, thermophilic bacteria were isolated from milk samples obtained from a milk processing plant to investigate the potential of different isolates to convert nitrate to nitrite. Eight bacteria species were identified through 16s rDNA gene sequencing (Bacillus licheniformis, Bacillus aerius, Bacillus subtilis, Bacillus sonorensis, Bacillus firmus, Enterococcus faecium, Geobacillus stearothermophilus and Macrococcus caseolyticus). Most bacteria isolated were able to convert nitrate to nitrite, and surprisingly in aerobic conditions. The most interesting species was G. stearothermophilus as its nitrate reducing capability was highly variable between different isolates. This could explain why there was no relationship reported between nitrite level in milk powder and high thermophile count.     

Whey fermentation by thermophilic lactic acid bacteria: evolution of carbohydrates and protein content

Whey, a by-product of the cheese industry usually disposed as waste, is a source of biological and functional valuable proteins. The aim of this work was to evaluate the potentiality of three lactic acid bacteria strains to design a starter culture for developing functional whey-based drinks. Fermentations were performed at 37 and 42 degrees C for 24h in reconstituted whey powder (RW). Carbohydrates, organic acids and amino acids concentrations during fermentation were evaluated by RP-HPLC. Proteolytic activity was measured by the o-phthaldialdehyde test and hydrolysis of whey proteins was analyzed by Tricine SDS-PAGE. The studied strains grew well (2-3log cfu/ml) independently of the temperature used. Streptococcus thermophilus CRL 804 consumed 12% of the initial lactose concentration and produced the highest amount of lactic acid (45 mmol/l) at 24h. Lactobacillus delbrueckii subsp. bulgaricus CRL 454 was the most proteolytic (91 microg Leu/ml) strain and released the branched chain amino acids Leu and Val. In contrast, Lactobacillus acidophilus CRL 636 and S. thermophilus CRL 804 consumed most of the amino acids present in whey. The studied strains were able to degrade the major whey proteins, alpha-lactalbumin being degraded in a greater extent (2.2-3.4-fold) than beta-lactoglobulin. Two starter cultures were evaluated for their metabolic and proteolytic activities in RW. Both cultures acidified and reduced the lactose content in whey in a greater extent than the strains alone. The amino acid release was higher (86 microg/ml) for the starter SLb (strains CRL 804+CRL 454) than for SLa (strains CRL 804+CRL 636, 37 microg/ml). Regarding alpha-lactalbumin and beta-lactoglobulin degradation, no differences were observed as compared to the values obtained with the single cultures. The starter culture SLb showed high potential to be used for developing fermented whey-based beverages.     

Analysis of the methionine biosynthetic pathway in the extremely thermophilic eubacterium Thermus thermophilus

Four DNA fragments that could rescue the mutations of four Met- mutants were cloned from Thermus thermophilus HB27 and their complete nucleotide sequences were determined. Two of the four fragments respectively contained the greater parts of the metF and metH genes, the predicted amino acid sequences of which showed identities of 30.8% and 32.7% with 5,10-methylenetetrahydrofolate reductase (EC 1.7.99.5) and vitamin B12-dependent homocysteine transmethylase (EC 2.1.1.13) of Escherichia coli. The other two DNA fragments, which overlapped one another, contained two open reading frames whose predicted amino acid sequences were respectively similar to those of O-acetylhomoserine sulfhydrylase (EC 4.2.99.10, the product of the MET17 gene) and homoserine O-acetyltransferase (EC 2.3.1.31, the product of the MET2 gene) of Saccharomyces cerevisiae. The metF, metH, MET2, and MET17 genes of T. thermophilus were disrupted by introducing the heat-stable kanamycin nucleotidyltransferase gene into the genome. Each transformant showed methionine auxotrophy. Both the MET2- and MET17-disrupted mutants could grow in a minimal medium containing homocysteine but not in the same medium containing succinylhomoserine or cystathionine. In contrast, the metF- and metH-disrupted mutants could not grow in the minimal medium containing homocysteine. These results suggest that in T. thermophilus, homoserine is directly converted to homocysteine via O-acetylhomoserine and that homocysteine is methylated to synthesize methionine.     

Antimicrobial effects of green tea polyphenols on thermophilic spore-forming bacteria

The inhibitory action of tea polyphenols towards the development and growth of bacterial spores was examined. Among the tested Bacillus bacteria, tea polyphenols showed antibacterial effects towards Bacillus stearothermophilus, which is a thermophilic spore-forming bacterium. The heat resistance of B. stearothermophilus spores was reduced by the addition of tea polyphenols. Clostridium thermoaceticum, an anaerobic spore-forming bacterium, also exhibited reduced heat resistance of its spores in the presence of tea polyphenols. (-)-Epigallocatechin gallate, which is the main component of tea polyphenols, showed strong activity against both B. stearothermophilus and C. thermoaceticum. The heat resistance of these bacterial spores was more rapidly decreased by the addition of tea polyphenols at high temperatures.     

堆肥中高温纤维素酶菌株的筛选及其酶性质研究

从堆肥中分离获得20株耐高温纤维素分解菌,经过复筛,HC-5菌株产酶活性最高。HC-5的生长特性表明,其最适pH值为7.0,最适温度为50℃。菌株HC-5的CMCase和β-glucosidase酶作用活性范围均在40℃～70℃,但最适作用温度分别为60℃和50℃;Avicelase酶作用活性范围在40℃～60℃,而在50℃时酶作用活性最高。HC-5所产的纤维素分解酶具有良好的热稳定性,但随贮温的增加,其热稳定性呈现下降的趋势。     

Production of anti-prion scFv-Fc fusion proteins by recombinant animal cells

We constructed a replication-defective retroviral vector plasmid for the expression of a single-chain antibody fragment (scFv), derived from a chicken anti-human prion protein monoclonal antibody, fused with the Fc region of human IgG1. CHO-K1 and NS-1 cells were transformed with the viral vector pseudotyped with vesicular stomatitis virus G protein (VSV-G), and scFv-Fc producer clones were established. Among the established clones, CHO-2A9 cells produced a large amount of the product with an antibody-like dimerized structure in serum-free culture that facilitated the purification of scFv-Fc. The scFv-Fc specifically recognized the epitope sequence of prion protein in solid-phase enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. The injection test into quails revealed that the scFv became more stable in vivo by fusion with the Fc region. The scFv-Fc will be a useful tool for the detection of mammalian prion proteins.     

比色法检测乳中掺加的动物胶原水解蛋白

采用比色法快速定量检测乳中掺加的动物胶原水解蛋白。首先通过比色方法测得羟脯氨酸的质量浓度，由测得的羟脯氨酸换算得出乳中掺加的动物胶原水解蛋白的量。此法快速、简便、重复性好、回收率高。     

高效液相色谱法检测牛乳中掺加的胶原水解蛋白

本研究的目的是建立采用高效液相色谱测定牛乳中掺加胶原水解蛋白的方法.其基本原理是通过高效液相色谱法测得羟脯氨酸的含量,由测得的羟脯氨酸含量换算得出乳中掺加的胶原水解蛋白的量.以symmetry-C18(150 mm ×3.9 mm,i.d.5 μm)为色谱柱,2,4-二硝基氟苯为衍生试剂,梯度洗脱.流动相A:浓度为0.05 mol/L乙酸钠缓冲溶液(pH=6.5,含10 mL/L四氢呋喃),流动相B:乙腈-水(体积比1:1).羟脯氨酸质量浓度在2～10 mg/L的范围内与峰面积之间具有良好的线性关系,回收率为93.47%～101.20%,检出限为2.5 mg/L.测定结果的相对标准偏差为1.62%.     

浓缩果汁生产厂中嗜酸耐热菌的跟踪检测

研究探索用美国KRUEGER实验室分析检测嗜酸耐热菌方法,探讨了不同厂家的化学试剂对检测结果的影响,采取改进了的双膜过滤测定法对浓缩果汁生产厂中不同地点的空气,水质取样9检测,该法加大了取样量,在常量取样未能检出的取样点检出的嗜酸耐热菌,弄清了该菌在浓缩果汁生产厂中的污染途径,对采取相应的防范措施提供了参考依据。     

Antihypertensive peptides from whey proteins fermented by lactic acid bacteria

Food Science and Biotechnology - In this study, whey proteins were fermented with 34 lactic acid bacteria for 48 h at 37 °C and their ability to inhibit angiotensin...     

酶解动物蛋白制备抗氧化多肽研究进展

动物蛋白资源丰富,酶法水解动物蛋白制备的抗氧化多肽具有多种生理机能,成为研究的热点。综述了酶解动物蛋白制备抗氧化多肽的研究现状,为提高畜禽副产品附加值提供了理论依据。     

DNA fingerprinting of thermophilic lactic acid bacteria using repetitive sequence-based polymerase chain reaction

DNA fingerprints of lactic acid bacteria were generated by polymerase chain reaction using a primer based on the repetitive elements found in the genome of Streptococcus pneumoniae (BOX-PCR). The method made it possible to identify 37 isolates from raw milk. industrial starters and yogurt. Differentiation at species, subspecies and strain level was possible for Lactobacillus delbrueckii subsp. lactis, Lb. delbrueckii subsp bulgaricus and Str. thermophilus. BOX-PCR was also applied to studying the strain composition of a starter culture and the direct detection of strains in commercial fermented milk.     

Potential of thermophilic amylolytic bacteria for growth in unconventional media: Potato peels

Extremozymes have gained high level of popularity for their industrial relevance. In this regard four thermophilic bacterial isolates were isolated from hot spring Chakwal in Pakistan. All the strains were screened for amylolytic activity by producing zone of clearance on starch agar plates. The bacterial isolates were cultivated employing potato peels as major energy source. All the four bacterial strains were Gram positive, motile, endospore formers and were positive for catalase and oxidase tests. Phylogenetic analysis based on 16S rRNA sequences confirmed that all the strains were belonged to the Bacillus licheniformis with 98–99% similarity under Accession numbers KF424263, KF424264, KF424265, and KF424266 for KA2, KA5, KA6, and KA9, respectively. The pH, temperature, oxygen requirement, and inocula size of the medium were optimized, which could yield amylase up to 0.61 U while growing in potato peels based media. Besides being thermostable, the enzymes have a working pH range of 5–9. The isolate KA2 showed maximum threefold purification, and percentage yield was estimated to be 93.06% as compared to crude enzyme. They yielded enough protein suggesting their potential in industrial applications in unconventional and economical substrate.

Practical applications

Amylases produced in this study could be utilized in starch hydrolysis for the production bioethanol and can be employed in different industries like food, paper, and detergent for various purposes. In addition, production of efficient amylases resolves the waste management problem of agro‐industrial waste of potato peels.     

Technological properties of milks fermented with thermophilic lactic acid bacteria at suboptimal temperature

In the present work the synergistic relationship between different strains of Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus was studied at optimal (44 degrees C) and suboptimal temperatures (30 degrees C). Acidification, viscosity, whey syneresis, and bacterial concentration of the final product were evaluated on single-strain and mixed cultures after 24 h at 30 degrees C and 6 h at 44 degrees C. Three pairs of strains (LBB + CP2, LBP + CP2, and LBR + CP2) showed synergistic effect, which was reflected by the viscosity and syneresis of the coagulum. These results were more significant when cultures were incubated at 30 degrees C, reaching apparent viscosity values of 19 to 28 mPa x s. On the other hand, lactobacilli cultures enhanced the growth of two streptococci strains (CP2 and CP4). These results were confirmed by cultures of streptococci supplemented with supernatants of culture of lactobacilli. Those supernatants stimulate the viscosity produced by CP2 and CP4 strains and reduce the syneresis of all cultures of streptococci. Neither the increase of viscosity nor reduction of syneresis could be attributed to a decrease of pH.     

Microbial factories for the production of animal proteins

Recombinant DNA technology has recently been shown to offer a route to the microbial synthesis of significant amounts of animal, viral, and human proteins. In order to take advantage of this technology, a desired gene must first be obtained and cloned in a suitable host organism. The foreign gene must then be efficiently expressed. In many cases, the foreign gene product must then be isolated. Results obtained through cloning and expressing the chicken ovalbumin gene in microorganisms can be used to demonstrate both the power of this technology and its present limitations.     

芝麻香型白酒高温大曲嗜热功能菌的筛选与鉴定

以芝麻香型白酒高温大曲为材料,采用55℃高温分离培养,通过平板透明圈法初筛和三角瓶固态发酵物蛋白酶活力测定复筛,分离筛选嗜热功能细菌,并结合16S rRNA基因序列分析和系统发育分析进行菌株鉴定。结果表明,55℃条件下共分离获得85株嗜热细菌,其中M5中性蛋白酶活力为96.06 U/g,优于对照菌株。菌株M5经分子生物学鉴定为枯草芽孢杆菌枯草亚种(Bacillus subtilis subsp.Subtilis),该菌株可制成微生物强化菌剂,应用于芝麻香型白酒的高温制曲过程中。     

Targets and methods for the detection of processed animal proteins in animal feedstuffs

Bovine spongiform encephalopathy (BSE), or ‘mad cow disease’, is one of several transmissable spongiform encephalopathies (TSEs) known to affect certain mammals and is spread through the ingestion of infected animal tissue. It is believed that the inadvertent contamination of meat and bone meal (MBM) with infected animal tissue and the subsequent use of this material as a feed supplement contributed to the spread of the disease in cattle. As a result, the use of processed animal proteins (PAPs) in animal feeds is regulated in many parts of the world. Although feed testing is the only definitive means to certify compliance, regulatory compliance often relies solely on paper certification. Recently, rapid methods have become available that can be used by regulators to determine compliance during routine inspections. We describe a rapid, immunochromatographic strip test that can detect 0.1% MBM in animal feed. The test takes 15 min to perform and large numbers of samples can be screened for PAPs simultaneously.