吸附—交联法固定胰蛋白酶及在澄清啤酒中的应用

胰蛋白酶的固定化采用吸附－交联法。在单因素试验基础上，通过正交试验，获得了胰蛋白酶固定化的最佳条件：酶与载体体积比为2：1，吸附时间为10h,pH＝4．5，缓冲液浓度为0．05mol/L，戊二醛的质量分数为0．1％。此条件下制得的磁性酶活力回收在50％左右，相对酶活力达74％，该磁性酶对啤酒澄清防止冷浑浊有明显效果。     

酶法水解中国毛虾的研究(英文)

比较了4种常用蛋白酶对中国毛虾的酶解效果,筛选出胰蛋白酶和枯草杆菌蛋白酶为最佳用酶,并利用正交试验优化了水解条件,在各自最佳水解条件下采用先加胰蛋白酶再加枯草杆菌蛋白酶的酶解方式对中国毛虾进行联合双酶水解,水解度可达67 .83%。     

酶法水解中国毛虾的研究

比较了4种常用蛋白酶对中国毛虾的酶解效果，筛选出胰蛋白酶和枯草杆菌蛋白酶为最佳用酶，并利用正交试验优化了水解条件，在各自最佳水解条件下采用先加胰蛋白酶再加枯草杆菌蛋白酶的酶解方式对中国毛虾进行联合双酶水解，水解度可达67．83％。     

黑曲霉产β-葡萄糖苷酶工艺的神经网络优化

在单因素试验研究的基础上,采用Box-Behnken中心组合设计,利用JMP 7.0中的神经网络平台,在接种量、初始pH、发酵时间和装液量等4个方面对黑曲霉CICC 2475发酵产β-葡萄糖苷酶工艺条件进行优化,以期获得高酶活力的β-葡萄糖苷酶。结果表明:接种量约11.0%、初始pH值5.6、发酵时间130 h以及装液量70.0 mL条件下,黑曲霉发酵产β-葡萄糖苷酶的酶活力达到最大值118.73 U/mL,与单因素优化前61.01 U/mL相比,酶活力提高了48.61%。     

太平洋鳕仔鱼期消化酶活性的变化

用酶学分析法研究太平洋鳕仔鱼发育过程中胰蛋白酶、酸性蛋白酶、淀粉酶和脂肪酶活性的变化。结果表明:4种酶的比活力均在初孵时即可检出,但比活力变化趋势不同;胰蛋白酶比活力在仔鱼孵化初期较高,随后显著降低;酸性蛋白酶比活力在早期仔鱼阶段比较低,且较为稳定,从14日龄开始显著升高;淀粉酶比活力在3日龄即达到最大值,之后显著降低,并一直保持较低的比活力;脂肪酶的比活力一直较低,在5日龄达到最大值,之后比活力大幅度下降。仔鱼发育过程中,各种酶活性变化显著(p<0.05),太平洋鳕在仔鱼期已经可以消化蛋白质、脂肪和碳水化合物。     

酶解法提取鱼油的工艺参数优化

利用蛋白酶酶解法从黄鳍金枪鱼(Thunnus albacares)加工的下脚料———鱼头中提取鱼油。以鱼油提取率和感官特征为指标,通过正交优化实验设计,获得了胰蛋白酶提取鱼油的最佳酶解工艺参数:酶解温度45℃,酶添加量1.5%,料液质量比1∶1,酶解时间4 h,酶解pH 8。在该条件下,鱼油的提取率为4.34%,理化指标除过氧化值外均达到SC/T3502-2000的粗鱼油二级标准,多不饱和脂肪酸总含量高达38.47%,其中DHA和EPA的含量分别为23.63%和4.84%。     

微囊藻毒素对凡纳滨对虾免疫相关酶活力的影响

通过毒性注射实验,探究微囊藻毒素(MC-LR)对凡纳滨对虾(Litopenaeus Vannamei)肝脏免疫相关酶活力的影响。结果表明,注射MC-LR后,凡纳滨对虾肝胰脏组织的超氧化歧化酶、谷胱甘肽过氧化酶、过氧化物酶、酸性磷酸酶、碱性磷酸酶和溶菌酶的酶活力呈先增大后减小的变化趋势。超氧化歧化酶、谷胱甘肽过氧化酶和过氧化物酶活力在对虾注射MC-LR 4 h后达到峰值,比对照组分别增加了15.62%、18.76%和32.86%,随后酶活力逐渐减小;而酸性磷酸酶和碱性磷酸酶酶活力在对虾注射MC-LR 2 h后即显著增加,比对照组分别增加了25.02%和37.99%,随后酶活力逐渐减小;溶菌酶在对虾注射MC-LR后8 h达到最大值,比对照组增加了33.57%,随后酶活力逐渐减小。MC-LR能显著影响凡纳滨对虾的免疫相关酶活力(P0.05),引起凡纳滨对虾的应激反应和抑制它们的免疫相关酶活力。     

三角帆蚌肉酶解产物响应面优化制备及醒酒活性

【目的】研究三角帆蚌(Hyriopsis cumingii)肉酶解产物的解酒防醉作用。【方法】以乙醇脱氢酶(ADH)激活率为指标,在单因素试验基础上响应面优化三角帆蚌肉酶解工艺,制备三角帆蚌肉酶解产物(Enzymatic hydrolysate from soft tissue of Hyriopsis cumingii, EH),并测定EH对饮酒小鼠醉酒状态、血液中乙醇浓度和肝脏ADH活力的影响,以评价其醒酒活性。【结果】选用中性蛋白酶为实验用酶,三角帆蚌肉的最佳酶解条件为酶解时间2.0 h、酶解温度50.0℃、料液比1∶3、加酶量1 000 U/g,在此条件下ADH激活率为18.30%。小鼠摄入EH300 mg/kg时,饮酒小鼠醉酒只数少,酒后90 min血乙醇浓度已降至8.80 mmol/L,肝脏ADH活力上升至7.64 U/mg。【结论】三角帆蚌肉酶解产物能够显著激活肝脏ADH,降低血乙醇浓度,具有较好的醒酒功效和应用前景。     

菠萝天然酶活力测定方法及活力变化

以菠萝 (AnanascomosusMerr)为原料 ,应用改良的方法对菠萝天然酶活力进行测定 ,对菠萝酶活力变化进行了初步探讨。结果表明 :测定未成熟、成熟菠萝酶活力分别为 10 6 .10和 6 5 .85u/mL ;菠萝肉、皮、整果汁酶活力分别为 86 .98、5 0 .72、5 5 .2 1u/mL ;室温贮藏 ,酶活力取样时为 5 4.97u/mL ,第 4天达最高值 72 .78u/mL ,第 8天降至 4 3.6 6u/mL ;4℃贮藏 ,酶活力取样时为 5 4.97u/mL ,第 2天达最高值71.6 8u/mL ,第 4天降至 5 0 .12u/mL ,即不同温度下贮藏不同时间酶活力变化是先升高后降低     

波吉卵囊藻的固定化培养及利用

以褐藻酸钙为固定化载体,探讨了胶球直径、不同接种量、CaCl2溶液浓度、褐藻酸钠浓度等条件对波吉卵囊藻(Oocystisborgei)固定化培养及对养殖水体水质的影响。结果表明:波吉卵囊藻固定化培养的最好条件是微藻接种量为1×106 mL-1,CaCl2溶液的质量浓度为30g/L,褐藻酸钠溶液的质量浓度为20g/L。在此条件下制备的固定化藻珠,波吉卵囊藻的生物量从1 83×10-2 mg/粒增加到17. 46×10-2 mg/粒,增加了近10倍,生长率相对较高,证明了它的生理活性不会因固定化而受干扰。引入固定化的波吉卵囊藻不但可以降低养殖水体中氨氮、亚硝酸氮等有害因子的浓度,还能提高水中溶解氧含量,使水体环境长时间处于良好的动态平衡状态。     

生物采油解堵剂中产蛋白酶菌株的筛选及培养条件优化

为研究生物酶采油解堵剂中产蛋白酶菌株的初、复筛选及培养条件优化,从大庆原油样品中筛选菌种,通过水解酪素的透明圈实验及福林酚测蛋白酶酶活的方法进行菌株的初、复筛选;以蛋白酶酶活为优化指标,采用单因素实验对筛选的产蛋白酶菌株的培养基及培养条件进行优化,优化最适培养基:可溶性淀粉为15g/L,蛋白胨为20g/L,酵母膏为20g/L,NaCl为1.0g/L,CaCl2为0.02g/L,Na2HPO4为0.2g/L,NaH2PO4为0.1g/L;在初始pH为6.0、接种量为5%(体积分数)、温度为31℃、摇床转速为160r/min的条件下,培养72h后,菌株的蛋白酶酶活为551.0U/mL,为复筛选菌株的蛋白酶酶活的22.92倍,即为菌株生长繁殖及代谢的最佳条件,能够获得更高的蛋白酶酶活,有利于后续实验的进行.结果表明:菌株产蛋白酶对原油作用效果为发酵液表面张力从作用前的56.2mN/m降低到作用后的30.5mN/m,表面张力显著降低,还有降解降黏原油等效果,具有一定的研究价值.     

米曲霉中性蛋白酶特性的研究

研究了米曲霉中性蛋白酶的特性。结果表明,米曲霉中性蛋白酶的最佳提取方法为加入pH 7.2的缓冲液研匀,置恒温(30℃)培养箱中提取30 min,每10 min搅动1次,最后用4层纱布过滤。酶反应最适温度为55℃,最适pH值为7.2。NaCl和EDTA对其有一定的抑制作用。酶的热稳定性好,在45℃的温度下处理24 h后,酶活力仍有45.5%。米曲霉中性蛋白酶与培养物一起保存时,稳定性也较好,在4℃保存一个月后,酶活力几乎无变化。     

Isolation and characterization of a marine bacterium producing protease from Chukchi Sea, Arctic

A Gram negative bacterium Ar/W/b/75°25＇N/1 producing extracellular alkaline protease was isolated from surface water of latitude 75°25＇N, and longitude 162°25＇W in Chukchi sea, Arctic. The strain can grow at the temperature range from 7℃ to 30℃, and grow better at 30(℃. It can not grow at 40℃. Keeping certain salinity concentration in medium is necessary for cell growth. It grows well in medium containing salinity concentration from 0. 5 % to 10 % sodium chloride. Glucose, sucrose and soluble starch can be utilized by the strain, among which glucose is the optimal carbon source. Peptone is the optimal organic nitrogen source for cell growth and protease producing, and ammonium nitrate is the optimal inorganic nitrogen source.About 75.7% of total protease of the strain are extracellular enzyme. Optimal temperature for proteolytic activity is at 40℃. Protease of the strain keeps stable below 40℃, and shows high proteolytic activity within the pH range from 7 to 11.     

Properties of Klebsiella phage P13 and associated exopolysaccharide depolymerase

The bacteriophage P13 that infects Klebsiella serotype K13 contains a heat-stable depolymerase capable of effective degradation of exopolysaccharide(EPS) produced by this microorganism. In this study, the titer of phage P13, initially 2.0 × 107 pfu mL-1, was found increasing 20 min after infection and reached 5.0 × 109 pfu mL-1 in 60 min. Accordingly, the enzyme activity of depolymerase approached the maximum 60 min after infection. Treatment at 70℃ for 30 min inactivated all the phage, but retained over 90% of the depolymerase activity. Addition of acetone into the crude phage lysate led to precipitation of the protein, with a marked increase in bacterial EPS degradation activity and a rapid drop in the titer of phage. After partial purification by acetone precipitation and ultrafiltration centrifugation, the enzyme was separated from the phage particles, showing two components with enzyme activity on Q-Sepharose Fast Flow. The soluble enzyme had an optimum degradation activity at 60℃ and pH 6.5. Transmission electron microscopy demonstrated that the phage P13 particles were spherical with a diameter of 50 nm and a short stumpy tail. It was a double-strand DNA virus consisting of a nucleic acid molecule of 45976 bp. This work provides an efficient purification operation including thermal treatment and ultrafiltration centrifugation, to dissociate depolymerase from phage particles. The characterization of phage P13 and associated EPS depolymerase is beneficial for further application of this enzyme.     

青鳞鱼蛋白复合酶控制水解物功能特性的研究

研究了青鳞鱼蛋白复合酶控制水解产物的功能特性。结果表明,复合酶控制水解能够显著地改善水解物的溶解性、乳化活力及起泡性能。水解物在pH 2.0~10.0范围内其NSI为89.3%~98.5%,且在酸性范围NSI高于碱性范围;在酸性pH内水解物的乳化活力都高于112 m2/g,随着DH的增加,最大乳化活力出现的pH范围更宽广,但水解却降低了乳化稳定性。蛋白质质量分数为1%~5%时,FP随质量分数的增加而增加;泡沫形成后,未水解的蛋白质具有最高的稳定性,DH为8.4%的水解物具有最高的稳定性。     

Purification and characterization of iron-cofactored superoxide dismutase from Enteromorpha linza

A superoxide dismutase was purified from Enteromorpha linza using a simple and safe procedure, which comprised phosphate buffer extraction, ammonium sulphate precipitation, ion exchange chromatography on Q-sepharose column, and gel filtration chromatography on Superdex 200 10/300GL. The E. linza superoxide dismutase （E/SOD） was purified 103.6-fold, and a yield of 19.1% and a specific activity of 1 750 U/rag protein were obtained. The SDS-PAGE exhibited E/SOD a single band near 23 kDa and the gel filtration study showed E/SOD＇s molecular weight is near 46 kDa in nondenatured condition, indicating it＇s a homodimeric protein. E/SOD is an iron-cofactored superoxide dismutase （Fe-SOD） because it was inhibited by hydrogen peroxide, insensitive to potassium cyanide. The optimal temperature for its maximal enzyme activity was 35℃, and it still had 29.8% relative activity at 0℃, then E/SOD can be classified as a cold-adapted enzyme. E/SOD was stable when temperature was below 40℃ or the pH was within the range of 5 10. The first 11 N-terminal amino acids orE/SOD were ALELKAPPYEL, comparison of its N-terminal sequence with other Fe-SOD N-terminal sequences at the same position suggests it is possibly a chloroplastic Fe-SOD.     

Purification and characterization of an alkaline protease from <Emphasis Type="Italic">Micrococcus</Emphasis> sp. isolated from the South China Sea

Protease is wildly used in various fields, such as food, medicine, washing, leather, cosmetics and other industrial fields. In this study, an alkaline protease secreted by Micrococcus NH54PC02 isolated from the South China Sea was purified and characterized. The growth curve and enzyme activity curve indicated that the cell reached a maximum concentration at the 30th hour and the enzyme activity reached the maximum value at the 36th hour. The protease was purified with 3 steps involving ammonium sulfate precipitation, ion-exchange chromatography and hydrophobic chromatography with 8.22-fold increase in specific activity and 23.68% increase in the recovery. The molecular mass of the protease was estimated to be 25 kDa by SDS-PAGE analysis. The optimum temperature and pH for the protease activity were 50°C and pH 10.0, respectively. The protease showed a strong stability in a wide range of pH values ranging from 6.0–11.0, and maintained 90% enzyme activity in strong alkaline environment with pH 11.0. Inhibitor trials indicated that the protease might be serine protease. But it also possessed the characteristic of metalloprotease as it could be strongly inhibited by EDTA and strongly stimulated by Mn²⁺. Evaluation of matrix-assisted laser desorption ionization/time-of-flight MS (MALDI-TOF-TOF/MS) showed that the protease might belong to the peptidase S8 family.     

Screening and characterization of the alkaline protease isolated from PLI-1, a strain of Brevibacillus sp. collected from Indonesia’s hot springs

A total of 69 strains of thermophilic bacteria were isolated from water, soil and sediment samples from three Indonesia’s hot spring areas (Pantai cermin, Kalianda and Banyu wedang) by using Minimal Synthetic Medium (MSM). The extreme thermophile Brevibacillus sp. PLI-1 was found to produce extracellular thermophilic alkaline protease with optimal activity at 70℃ and pH 8.0-9.0. The molecular weight of the protease was estimated to be around 56 kD by SDS-PAGE. The maximum activity of the protease was 26.54 U mL-1. The protease activity did not decrease after 30 min and still retained more than 70% of relative activity after 60 min at 70℃ and pH 8.0. The ion Mg2+ was found to promote protease activity at both low and high concentrations, whereas Cu2+ and Zn2+ could almost completely inhibit the activity. Divalent cation chelator EDTA inhibited the enzyme activity by 55.06% ± 0.27%, while the inhibition caused by PMSF, Leupeptin, Pepstain A and Benzamidine were 66.78% ± 3.25%, 52.37% ± 0.25%, 62.47% ± 2.96% and 50.99% ± 0.24%, respectively. Based on these observations, the enzyme activity was conspicuously sensitive to the serine and cysteine protease inhibitors. All these results indicated that the protease isolated from the strain PLI-1 was a thermophilic protease and had a high-temperature stability and a pH stability.     

Purification and Characterization of a Novel Lipase from Antarctic Krill

Lipase from Antarctic krill,with a molecular weight of 71.27kDa,was purified with ammonium sulfate precipitation and a series of chromatographic separations over ion exchange(DEAE)and gel filtration columns(Sephacryl S-100),resulting in 5.2%recovery with a 22.4-fold purification ratio.The optimal pH and temperature for enzyme activity were 8.0 and 45℃,respectively.Purified lipase had Km and Vmax values of 3.27mmolL−1 and 2.4Umg−1,respectively,using p-nitrophenyl laurate as the substrate.Lipase activity was enhanced by adding Ca2+and Mg2+ions in the concentration ranges of 0–0.5mmolL−1 and 0–0.3mmolL−1,respectively,while the activity was inhibited by a further increase in these ion concentrations.Fe3+and Cu2+ions showed obvious inhibitory effects on enzyme activity,and the inhibition rates were 71.8%and 53.3%when the ion concentrations were 0.5mmolL−1.