海因连续水解生成海因酸及其动力学研究

在碱性条件下,以海因法连续水解生成甘氨酸的方法是目前最环保的工艺。实验探究海因法的第一步反应,采用的反应器为管式反应器,以海因、氢氧化钠和水为原料,进行试验,得到最适反应条件为温度110℃,氢氧化钠与海因摩尔比为1.6∶1,停留时间25 min,使得海因转化率达到了99.7%。同时本文建立了海因水解的动力学模型,确定该水解反应为一级反应,并求得反应活化能E为2.9307×10^4J·mol^-1,指前因子A0为7.92×10^2。

海因消旋酶用于制备D-苯丙氨酸的工艺研究

研究酶法制备D-苯丙氨酸的合成工艺,在原有苄基海因水解工艺的基础上增加了海因消旋酶的投入,在转化率相同的情况下,反应温度可以下降25℃。通过优化工艺条件,确定了海因消旋酶的最佳反应温度。通过对比,增加海因消旋酶,可以极大地降低转化温度,有助于海因酶酶活的稳定,也有利于降低生产的能耗。该方法绿色环保,对环境污染小,适合工业化生产。

单扫描极谱法测定海因类化合物

提出了测定海因类化合物的新方法，其线性范围为１．０×１０－５～８．０×１０－５ｍｏｌ／Ｌ，用于海因类化合物的测定，得到了满意结果。

A New κ-Carrageenase Cgk S from Marine Bacterium Shewanella sp. Kz7

A new κ-carrageenase gene cgkS was cloned from marine bacterium Shewanella sp. Kz7 by using degenerate and site-finding PCR. The gene was comprised of an open reading frame of 1224bp, encoding 407 amino acid residues, with a signal peptide of 24 residues. Based on the deduced amino acid sequence, the κ-carrageenase CgkS was classified into the Glycoside Hy- drolase family 16. The cgkS gene was expressed in Escherichia coli, and the recombinant enzyme was purified to homogeneity with a specific activity of 716.8 U mg-1 and a yield of 69%. Recombinant CgkS was most active at 45 ℃ and pH 8.0. It was stable at pH 6.0-9.0 and below 30℃. The enzyme did not require NaCl for activity, although its activity was enhanced by NaCI. CgkS degraded κ-carrageenan in an endo-fashion releasing tetrasaccharides and disaccharides as main hydrolysis products.

Mechanisms and factors affecting the adsorption of sodium alginate onto modified clays

Algal organic materials （AOMs） are one critical factor affecting the efficiency of modified clays used for the mitigation of harrnful algal blooms （HABs）. This study was conducted to develop a deeper understanding of the mechanisms and factors affecting the adsorption of AOMs onto modified clays. Sodium alginate （polysaccharide） and kaolinite modified with polyaluminium chloride （PAC1） were used as AOMs and modified clay model substances, respectively, and the effects of modifier dosage, contact time, solution pH and ionic strength were investigated through batch adsorption experiments. Kinetics revealed that the alginate adsorption rate was described well by a pseudo-second order model. PACl effectively enhanced the adsorption capacity of kaolinite and increased the adsorption rate, and the optimum additive amount of PACl was 5%. The experimental data fitted both the Freundlich and Langmuir adsorption equations well. The adsorption thermodynamics for alginate onto modified clays suggests that alginate adsorption is a spontaneous process. The adsorption of alginate onto modified clays was highly dependent on pH, with a decrease in adsorption observed with increased pH to 9.48, but the opposite was true above pH 9.48. Finally, adsorption increased with increasing ionic strength.

Detectingβ-thalassaemia mutations from a single cell by PEP and RDB

Objective :To evaluate the possibility of the technology involving PEP and RDB for detectingβ-thalassaemia multipoint mutations from a single cell simultaneously. Methods: A set of allele specific oligonucleotide (ASO) probes used for detecting 8 familiarβ-thalassaemia mutations (CD41-42. IVS-Ⅱ-654, CD17, TATA box nt-28, CD71-72, TATA box nt-29, CD26, IVS-Ⅰ-5) were immobilized on a strip of nylon membrane. The genome of a individual cell was amplified by primer extension preamplification (PEP) with the mixture of 15-base random oligonucleotides. The aliquots from PEP were used to amplify the objective gene fractions ofβ-thalassaemia gene by nested or semi-nested PCR. The membrane was hybridized with the final amplified products and then treated with Streptavidin-HRP and color development. Results:Totally 30 lymphocytes were picked up from blood samples of 1 healthy female and 4 patients with knownβ-thalassaemia mutations respectively. Each single lymphocyte was lysed in the proteinase K buffer. The amplification efficacy was 94. 0% and alle drop-out (ADO) rate was 8. 0%. Revert dot blot (RDB) was applied to the final amplified products from the 5 participants. The results of diagnosis were the same to the expected, and their genotypes were N/N, CD17(A→T)/N, IVS-Ⅱ-654(C→T)/CD17(A→T), CD41-42(-CTTT)/N and TATA box nt-28(A→G)/N, respectively. Conclusion: The technology involving PEP and RDB could detect multipleβ-thalassaemia mutations from a single cell simultaneously, and the research provides experimental evidences for the feasibility of applying PEP and DNA array technology to screening multiple genetic mutations from a single cell, and will be applied to preimplantation genetic diagnosis and non-invasive prenatal diagnosis forβ-thalassaemia.

Cloning and nucleotide sequence of D-hydantoinase gene of marine polyphosphate-accumulating bacterium, Halomonas sp. YSR-3

Hydantoinase is involved in the production of optically pure amino acids from racemic 5-mono-substituted hydantoins. We measured the D-hydantoinase activity in marine Halomonas sp. YSR-3 and amplified the D-hydantoinase gene by PCR. The gene was inserted into vector pGM-T and transformed into E. coli TOP10. The positive transformants with the D-hydantoinase gene were sequenced. The sequenced fragment comprises 1 510 base pairs. The D-hydantoinase gene from YSR-3 is 77% similar to that from Pseudomonas entornophila L4 by searching against the NCBI databse. The protein product of the YSR-3 D-hydantoinase gene is 75%, 73%, and 70% similar to those from Pseudomonasfluorescens Pf-5, Marinornonas sp. MED121, and Burkholderia vietnamiensis G4, respectively. The difference of the D-hydantoinase gene between marine Halornonas sp. YSR-3 and other terrestrial organisms is distinct.

Molecular Characterization of Sulfate-Reducing Bacteria Community in Surface Sediments from the Adjacent Area of Changjiang Estuary

Sulfate-reducing bacteria(SRB),which obtain energy from dissimilatory sulfate reduction,play a vital role in the carbon and sulfur cycles.The dissimilatory sulfite reductase(Dsr),catalyzing the last step in the sulfate reduction pathway,has been found in all known SRB that have been tested so far.In this study,the diversity of SRB was investigated in the surface sediments from the adjacent area of Changjiang Estuary by PCR amplification,cloning and sequencing of the dissimilatory sulfite reductase beta subunit gene(dsr B).Based on dsr B clone libraries constructed in this study,diversified SRB were found,represented by 173 unique OTUs.Certain cloned sequences were associated with Desulfobacteraceae,Desulfobulbaceae,and a large fraction(60%) of novel sequences that have deeply branched groups in the dsr B tree,indicating that novel SRB inhabit the surface sediments.In addition,correlations of the SRB assemblages with environmental factors were analyzed by the linear model-based redundancy analysis(RDA).The result revealed that temperature,salinity and the content of TOC were most closely correlated with the SRB communities.More information on SRB community was obtained by applying the utility of Uni Frac to published dsr B gene sequences from this study and other 9 different kinds of marine environments.The results demonstrated that there were highly similar SRB genotypes in the marine and estuarine sediments,and that geographic positions and environmental factors influenced the SRB community distribution.

Research and development of alginate-based raft forming system

Formulations of Gaviscon based on novel alginate raft forming system have been developed to prevent gastroesophageal reflux disease（GERD） symptoms. Alginate gel becomes buoyant and CO2-aerated as soon as Gaviscon comes into contact with gastric contents. This system can be characterized by several indicators, such as strength or coherence, volume, buoyancy, reflux resistance and resilience. In addition, the strength of the raft is influenced by many factors, such as molecular weight and the ratio of D-mannuronic and L-guluronic acid residues（M/G）, which are intrinsic factors in this formulation. Besides, there are several extrinsic factors, such as the presence of specific cations, the amount of carbon dioxide generated and entrapped in the raft. This review focused on how to evaluate the raft, introduced some factors that impact the raft and summarized the degradation of alginate and future advance.

Biomolecular characterization, identification, enzyme activities of molds and physiological changes in sweet potatoes(Ipomea batatas) stored under controlled atmospheric conditions

Microbial attacks during storage are one of the primary causes of product deterioration, and can limit the process of prolonging the shelf-life of harvested food. In this study, sweet potatoes were stored at temperatures of 13, 21, and 29 ℃ for 4 weeks. Samples were collected during storage and plated on potato dextrose agar, from which axenic mold cultures were obtained and identified using 26S rRNA gene sequences. Physiological changes of potato tubers were assessed with respect to pathogenicity, enzyme activity, and atmospheric storage conditions. Six fungal species were identified, namely Penicillium chrysogenum （P. rubens）, P. brevicompactum, Mucor circinelloides, C/adosporium cladosporiodes, P. expansum, and P. crustosum. The following fungal isolates, namely P. expansum, P. brevicompactum, and Rhizopus oryzae, were recovered from the re-infected samples and selected according to their levels of enzyme activity. This study revealed high levels of activity for cellulase and pectinase, which were most notable during the initial three days of testing, and were followed by a steady decrease （P〈0.05）. Polygalacturonase activity was prominent with values ranging from 12.64 to 56.79 U/mg （P. expansum） and 18.36 to 79.01 U/mg （P. brevicompactum）. Spoilage was obvious in the control group, which had a 100% decay at the end of the experimental period compared with samples treated with iprodione and sodium hypochlorite, in which the decay rates were 5% and 55%, respectively. The data for the iprodione- and sodium hypochlorite-treated samples at the end of the 3-month storage period showed that they were significantly different （P=0.041）, with the sodium hypochlorite-treated samples producing twice the rate of infection compared to the iprodione-treated samples. The comparative rate of the pro- gression of decay in the treated samples can be expressed as iprodione〈sodium hypochlorite〈control. This study demonstrates that sweet potato tissue damage is due to the activities of microbial enzymes and, in particular, the pectinases of the organisms isolated from the infected potato tissues, and suggests the advantages of utilizing iprodione as a curing agent for potato tubers before storage.