Effect of thermal pretreatment of raw soymilk on the gel strength and microstructure of tofu induced by microbial transglutaminase

The influence of thermal pretreatment of raw soymilk on the gel hardness and microstructure of tofu, induced by microbial transglutaminase (MTGase), was investigated in this paper. Modulated differential scanning calorimetry analysis showed that individual proteins in soymilk were to a various extent denatured by different thermal pretreatments. The viscosity of the soymilk and the gel hardness of MTGase-induced tofu were more highly related with the heating rate (up to 90 °C) than the mode of heating. At any enzyme concentration of MTGase, the tofus prepared from soymilk heated at 75 °C for 10 or 30 min showed highest gel hardness among all tested ones (P?0.05). Scanning electron microscopy analysis indicated that the microstructure of the tofu from soymilk heated at 75 °C for 30 min had a unique coral-like structure, much more continuous and homogenous than that from soymilk at 95 °C for 5 min. These results confirmed that the appropriate heat pretreatment (e.g. in the present, at 75 °C for 10-30 min) remarkably improved the gel strength of tofu by means of MTGase, and strengthened the tofu gel structure.     

Formation and rheological properties of ‘cold-set’ tofu induced by microbial transglutaminase

Microbial transglutaminase (MTGase) has been shown to effectively induce soymilk with a certain level of solid content to form filled tofu. The gel formation of this kind of tofu and the influence of reaction parameters on the properties of formed tofu were investigated by dynamic oscillatory and/or large-strain rheological measurements. The gelation process and the development of the mechanical moduli (especially the storage modulus, G′) of this kind of tofu were highly dependent upon the incubation temperature. Textural property analysis (TPA) results showed that many TPA parameters of this kind of tofu, including gel hardness, gumminess, springiness and cohesiveness, were also affected by the applied enzyme amount, the pH of soymilk and the presence of NaCl, especially for gel hardness. In addition, the additional heating and cooling treatment could significantly improve the gel strength of tofu, induced by MTGase at lower temperatures (e.g. 25 and 37 °C). These results suggested that a new kind of tofu with good quality could be produced using the enzymatic cross-linking technique, by the combination with the thermal treatment.     

Biodiversity and characterization of aerobic spore-forming bacteria in surimi seafood products

The microbial quality and safety of surimi seafood products was assessed by studying the prevalence and biodiversity of aerobic spore-forming bacteria at the beginning and end of shelf life in 100 surimi samples. Low levels of total flora and sporulated flora were numerated at the beginning of storage, however, residual spores were detected in the majority of samples during storage. Furthermore, for 34 samples, total flora counts > 10⁴ CFU/g were observed at the end of shelf life which could lead to non-compliance with good practice recommendations or product spoilage. In total, 460 strains were isolated, fingerprinted by M13-PCR and grouped into 98 different clusters. Representative strains were then identified at the species level via 16S rRNA gene sequencing. Overall, dominant species belonged to Bacillus simplex, Bacillus subtilis and Bacillus licheniformis; while B. simplex, B. subtilis as well as Sporosarcina aquimarina were clearly the dominant species found in samples with higher total flora counts. Amylolytic and proteolytic activities were very frequent amongst tested strains (80 and 92.5%, respectively). Heat resistance parameters of 4 strains in a surimi-based medium were determined. B. simplex and B. subtilis strains were the most heat resistant (δ_96 °C = 27.6 and 23.3 min and z_T = 8.6 and 7.9, respectively) which can explain their dominance in surimi samples exhibiting higher microbial counts. The heat resistance data obtained can now be used to model thermal destruction of strains using predictive microbiology tools (Sym’Previus).     

Sphingomyelinase C from Streptomyces sp. A9107: Unusual primary structure for bacterial sphingomyelinase C

A sphingomyelinase C (SMase) was identified in the culture supernatant of Streptomyces sp. A9107 (S-SMase). Although S-SMase seems to be a typical bacterial SMase, the primary structure of S-SMase was unusual for known bacterial SMase. The gene was functionally overexpressed in the culture medium of recombinant Rhodococcus erythropolis.     

Purification and characterization of transglutaminase from a newly isolated Streptomyces hygroscopicus

Transglutaminase (TGase, EC 2.3.2.13) from a Streptomyces hygroscopicus strain isolated from soil was purified from culture broth by ethanol precipitation, followed by successive chromatographies on CM-cellulose and Sephadex G-75 columns with a yield and purification-fold of 21.1% and 30%, respectively. The enzyme’s molecular weight was estimated as 38,000 Da by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified microbial transglutaminase (MTG) exhibited optimum activity at 37–45 °C and in a range of pH 6.0–7.0 for hydroxamate formation from N-carboxybenzoyl-l-glutaminyl-glycine and hydroxylamine. The enzyme was not stable above 50 °C and was stable within a pH range of 5.0–8.0 at lower temperature. The MTG was not inhibited by Ca²⁺ and ethylenediaminetetraacetic acid, suggesting it was calcium-independent. Purified MTG was strongly inactivated by 5,5′-dithiobis (2-nitrobenzoic acid), Cu²⁺, Zn²⁺, Pb²⁺, and Hg²⁺, suggesting that this enzyme could possess a thiol group at the active site. The MTG stability was strongly affected by ethanol concentration. The enzyme activity was slightly elevated at a lower concentration of ethanol at 25 °C.     

裂殖壶菌发酵产DHA油脂的生产工艺优化

采用磷酸香草醛法实时监测裂殖壶菌发酵产DHA油脂的积累情况,对裂殖壶菌的基础发酵工艺进行了优化。得到裂殖壶菌生长和油脂积累的最佳培养基配方为:葡萄糖30 g/L,玉米浆粉6 g/L,蛋白胨4 g/L,硝酸钠3.6~3.9 g/L,海水晶15 g/L;在50 L的发酵罐中采用后期流加一定量的葡萄糖提高碳氮比来提高油脂积累外,通过流加3.0 g/L的大豆油来刺激菌体生长,最终经过72 h的流加培养,菌体湿重达到200 g/L,总油脂含量达到60%以上,油脂脂肪酸组成中的DHA含量占22%左右。     

Complete detoxification of tris(1,3-dichloro-2-propyl) phosphate by mixed two bacteria, Sphingobium sp. strain TCM1 and Arthrobacter sp. strain PY1

Tris(1,3-dichloro-2-propyl) phosphate (TDCPP), a flame retardant, is regarded as a potentially toxic and persistent environmental contaminant. We previously isolated a TDCPP-degrading bacterium, Sphingobium sp. strain TCM1, which, however, produced a toxic metabolite: 1,3-dichloro-2-propanol (1,3-DCP). This study was undertaken to develop a technique for complete TDCPP detoxification using strain TCM1 with a 1,3-DCP-degrading bacterium, Arthrobacter sp. strain PY1. For efficient detoxification, we designed a resting cell system and examined the effect of freezing and lyophilization treatments for preparation of their resting cells. Results show that treatments had no marked adverse effect on their activities. The TDCPP dephosphorylation by TCM1 resting cells was optimal at 30°C and pH 8.5. Also, 1,3-DCP dehalogenation by strain PY1 resting cells was optimal at 35°C and pH 9.5. Under those respective conditions, the activities were 2.48 μmol h^− 1·OD₆₆₀^− 1 for TDCPP and 0.95 μmol h^− 1·OD₆₆₀^− 1 for 1,3-DCP. Based on these results, we set the reaction temperature to 30°C and pH to 9.0. Then we examined the detoxification of 50 μM TDCPP using mixed resting cells at a final OD₆₆₀ of 0.05 for strain TCM1 and 0.2 for strain PY1. In these conditions, TDCPP was eliminated after 1 h, but some of the resulting 1,3-DCP remained at a constant level. The increase in strain PY1 cells to a final OD₆₆₀ of 4.0 decreased the TDCPP dephosphorylation rate of strain TCM1 cells but achieved complete detoxification of TDCPP during 12 h of reaction.     

Enzymatic properties of transglutaminase produced by a new strain of Bacillus circulans BL32 and its action over food proteins

The aim of this study was to physicochemically characterize transglutaminase (TGase) from Bacillus circulans BL32, a strain recently isolated from the Amazon basin region, for its application in food systems. The effects of pH and temperature on the enzyme activity were determined by Central Composite Rotatable Design (CCRD), with maximal TGase activities obtained for pH between 5.7 and 8.7 and temperatures of 25-45 °C. This microbial TGase showed to be remarkably stable: over 90% of its activity was retained after 120 min of incubation at 50 °C. The Ca²⁺ and Mg²⁺ cations enhanced enzyme activity and its thermal stability when in concentrations of up to 2 and 1 mol L⁻¹, respectively. Casein, isolated soy protein, and hydrolysed animal protein were treated with this TGase. The decrease in the amount of free amino groups, especially for casein, showed the cross-linking of protein catalysed by this enzyme, while the emulsifying properties of these proteins were improved with treatment. These results suggest that this microbial TGase has a good potential to be used in food and other industrial applications.     

Purification and characterization of polyphenol oxidase from Ferula sp.

Polyphenol oxidase (PPO) of several Ferula sp. was extracted and purified through (NH₄)₂SO₄ precipitation, dialysis, and gel filtration chromatography. Leaf and stem extracts were used for the determination of enzyme properties. Optimum conditions, for pH, temperature, and ionic strength were determined. The best substrates of PPO were catechol for leaf and (−) epicatechin for stem samples. Optimum pH and temperature were determined. K_M and V_max values were 2.34 × 10⁻³ M and 8541 EU/ml for catechol, and 2.89 × 10⁻³ M and 5308 EU/ml for (−) epicatechin. The most effective inhibitor was sodium diethyl dithiocarbamate for leaf samples and sodium metabisulphite for stem samples. Both inhibitors indicated competitive reactions. PPO showed irreversible denaturation after 40 min at 60 °C.     

一株丝孢酵母属菌株发酵产油脂的研究

通过摇瓶发酵,比较丝孢酵母属菌株JM-B在不同营养和发酵条件下的产油脂情况。以菌体生物量、油脂产量、油脂含量及油脂系数为指标,优化碳源质量浓度、氮源组成与质量浓度、磷酸盐质量浓度等发酵培养基组成,优化接种量、发酵温度、摇床转速、发酵时间等发酵条件,利用气相色谱法测定油脂脂肪酸组成。结果表明:最适培养基组成为葡萄糖100 g/L,硫酸铵1.0 g/L,酵母膏10 g/L,磷酸二氢钾4 g/L,p H自然;最适发酵条件为发酵温度25℃,摇床转速160 r/min,种子液种龄36 h,接种量10%(以发酵液体积计),发酵时间144 h。在最适培养条件下,得到的菌体生物量和油脂产量最高,分别为16.14、6.64 g/L,油脂产量较优化前提高了161.4%,油脂中饱和脂肪酸含量比优化前降低了28.1%。可以认为丝孢酵母属菌株JM-B是一株很有开发潜力的产油脂菌株。     

Production and characterization of ectoine by Marinococcus sp. ECT1 isolated from a high-salinity environment

A halophilic bacterium isolated from a salt environment in southern Taiwan was identified as a Marinococcus sp. ECT1. This bacterium could synthesize and accumulate intracellular ectoine as a compatible solute capable of resisting osmotic stress in a hyper-osmotic environment. This study also developed a semi-synthesized medium (YAMS medium), capable of facilitating the growth of this Marinococcus sp. ECT1 with 600 mg/L crude ectoine production. Moreover, Marinococcus sp. ECT1 was grown on YAMS medium containing different initial yeast extract concentrations (C(YE)) (0 to 60 g/L) to demonstrate how C(YE) affects crude ectoine production. While the maximum cell concentration was increased by 23-fold when the C(YE) was 40 g/L, the maximum crude ectoine production reached 2.5 g/L when C(YE) was 40 g/L. In addition to demonstrating the success of the fermentation strategy of ectoine in increasing the production and production yield, experimental results further demonstrated that the fermentation medium of ectoine is highly promising for commercialization. Furthermore, the molecular weight and chemical structure of ectoine were identified and characterized by FAB-MS and (1)H-NMR.     

Production and purification of antioxidant peptides from a mungbean meal hydrolysate by Virgibacillus sp. SK37 proteinase

Antioxidant peptides of mungbean meal hydrolysed by Virgibacillus sp. SK37 proteinases (VH), Alcalase (AH) and Neutrase (NH) were investigated. The antioxidant activities based on 2,2′-azinobis (3-ethyl-benzothiazoline-6-sulphonate) (ABTS) radical-scavenging, ferric-reducing antioxidant power (FRAP) and metal chelation of VH were comparable to those of NH. VH was purified using ultrafiltration, ion exchange and gel filtration chromatography. The purified peptides (F37) from VH, which had the highest specific antioxidant activity, consisted of four peptides containing an arginine residue at their C-termini. In addition, the ABTS radical-scavenging activity of the purified peptides (F42) at 0.148 mg/ml was comparable to that of 1 mM of butylated hydroxytoluene (BHT). These two fractions were stable over a wide pH (4–10) and temperature (25–121 °C) range. Virgibacillus sp. SK37 proteinase is a potential processing-aid for the production of a mungbean meal hydrolyzate with antioxidant properties.     

Ammonium removal by Agrobacterium sp. LAD9 capable of heterotrophic nitrification-aerobic denitrification

Characteristics of ammonium removal by a newly isolated heterotrophic nitrification-aerobic denitrification bacterium Agrobacterium sp. LAD9 were systematically investigated. Succinate and acetate were found to be the most favorable carbon sources for LAD9. Response surface methodology (RSM) analysis demonstrated that maximum removal of ammonium occurred under the conditions with an initial pH of 8.46, C/N ratio of 8.28, temperature of 27.9°C and shaking speed of 150rpm, where temperature and shaking speed produced the largest effect. Further nitrogen balance analysis revealed that 50.1% of nitrogen was removed as gas products and 40.8% was converted to the biomass. Moreover, the occurrence of aerobic denitrification was evidenced by the utilization of nitrite and nitrate as nitrogen sources, and the successful amplifications of membrane bound nitrate reductase and cytochrome cd(1) nitrite reductase genes from strain LAD9. Thus, the nitrogen removal in strain LAD9 was speculated to comply with the mechanism of heterotrophic nitrification coupled with aerobic denitrification (NH(4)(+)-NH(2)OH-NO(2)(-)-N(2)O-N(2)), in which also accompanied with the mutual transformation of nitrite and nitrate. The findings can help in applying appropriate controls over operational parameters in systems involving the use of this kind of strain.     

红曲霉发酵不同底物产色素的研究进展

红曲色素是由红曲霉发酵产生的天然色素。传统上红曲霉都是发酵大米生产红曲色素,近些年,随着研究的深入,小米、玉米、木薯、菠萝蜜籽等谷物及农工业废弃物都被用于红曲色素生产。文章简述了红曲色素的性质,对红曲霉发酵不同底物生产色素的研究进行了综述。     

Complete detoxification of tris(2-chloroethyl) phosphate by two bacterial strains: Sphingobium sp. strain TCM1 and Xanthobacter autotrophicus strain GJ10

Tris(2-chloroethyl) phosphate (TCEP), a flame retardant, is recently regarded as a potentially toxic and persistent environmental contaminant. We previously isolated TCEP-degrading bacterium, Sphingobium sp. strain TCM1, which, however, produced a toxic metabolite: 2-chloroethanol (2-CE). This study was undertaken to develop a detoxification technique of TCEP using strain TCM1 with a 2-CE-degrading bacterium: Xanthobacter autotrophicus strain GJ10. TCEP degradation by strain TCM1-resting cells was thermally stable for 30 min at 30 °C. It was optimal at 30 °C and at pH 8.5. In the optimum condition, TCM1 cells up to a final cell density of 0.8 at OD(660) in the reaction mixture were unable to hydrolyze the phosphotriester bonds of 10 μM TCEP completely. The addition of 50 μM Co(2+) to reaction mixture enhanced the hydrolysis and caused the complete hydrolysis at the cell density of 0.8. Strain GJ10 resting cells degraded 2-CE only slightly, which might be attributable to lack of coenzyme regeneration of enzymes involved in the degradation. In contrast, the growing cells degraded approximately 180 μM of 2-CE within 24 h. Based on these results, we designed a two-step TCEP detoxification reaction consisting of TCEP hydrolysis to 2-CE by strain TCM1-resting cells and the following degradation of the resulting 2-CE by strain GJ10-growing cells. The combined reaction completely detoxified 10 μM TCEP, and thus opens a way to microbial detoxification of the potential toxic, persistent organophosphorus compound.     

轮枝霉(Diasporangiumsp.)产生EPA、AA发酵条件的初步研究

对本实验室分离的轮枝霉 (Diasporangiumsp )发酵产生含EPA、AA等多不饱和脂肪酸的条件 ,如碳源种类、氮源种类、pH、菌种的接种量和种龄、培养温度、通气量进行了研究 ,摇瓶发酵结果表明 ,以玉米粉和葡萄糖混合作为碳源能获得最高的不饱和脂肪酸产量 ;有机氮源有利于菌丝不饱和脂肪酸的产生 ;低温有利于菌丝中长链不饱和脂肪酸的产生 ,发酵接种量宜大 ;pH对菌丝生物量、EPA和AA的含量均有显著的影响。发酵培养 6d后菌丝中的不饱和脂肪酸含量达到最大值。在初步优化培养条件下 ,EPA、AA分别达到 2 5 3 7mg/L和 5 49 8mg/L。