ENZYMATIC CHARACTERISTICS OF TRYPSIN FROM PYLORIC CECA OF SPOTTED MACKEREL (SCOMBER AUSTRALASICUS)

Trypsin was purified from the pyloric ceca of spotted mackerel (Scomber australasicus) by gel filtration on Sephacryl S‐200 and Sephadex G‐50. The purification and yield were 20‐fold and 81%, respectively, as compared to those in the starting crude extract. Final enzyme preparation was nearly homogeneous in sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and the molecular weight of the enzyme was estimated to be 24,000 Da by SDS–PAGE. The trypsin was stable at pH 5–11 for 30 min at 30C, and its maximal activity against N^α‐p‐tosyl‐L‐arginine methyl ester was pH 8.0. Trypsin was heat‐stable up to about 50C for 15 min at pH 8.0. Optimum temperature of the trypsin enzyme was 60C. The enzyme was stabilized by calcium ion. The purified trypsin was strongly inhibited by serine protease inhibitors such as N‐p‐tosyl‐L‐lysine chloromethyl ketone and soybean trypsin inhibitor, suggesting that it is a trypsin‐like serine protease. N‐Terminal amino acid sequence of spotted mackerel trypsin was IVGGYECTAHSQPHQVSLNS.     

斯里兰卡蓝宝石和缅甸蓝宝石宝石学特征对比研究

运用显微观察等多种测试手段，在前人研究成果的基础上对斯里兰卡蓝宝石和缅甸蓝宝石的成因、产状及其宝石学特征进行了详细的对比研究。初步探讨了两地蓝宝石在宝石学特征方面的差异性，为鉴别斯里兰卡蓝宝石和缅甸蓝宝石提供了重要的实践依据。     

PURIFICATION AND PROPERTIES OF PHOSPHOLIPASE A2 ISOZYMES FROM PYLORIC CECA OF THE STARFISH (ASTERINA PECTINIFERA)

Phospholipase A₂ isozyme II (PLA₂ II), which showed different mobility on native PAGE from that of the PLA₂ isozyme I (PLA₂ 1) isolated previously, was purified from pyloric ceca of the starfish (Asterina pectinifera). The PLA₂ II mainly released oleic acid from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine. N-terminal amino acid sequence of the PLA₂ II was SVYQF. Temperature and pH optima of the PLA₂ II were at around 50C and pH 9.0, respectively, and the enzyme activity was enhanced by sodium deoxycholate and 1 mM or higher concentration of Ca²⁺. The PLA₂ II did not show fatty acid specificity for hydrolysis of phosphatidylcholine (PC). Specific activity of the PLA² II was about 10 times higher than that of commercially available porcine pancreatic PLA₂. The PLA₂ II hydrolyzed PC more effectively than phosphatidylethanolamine. These characteristics of the PLA₂ II were the same as those of the PLA₂ I.     

香菇谷氨酰胺转氨酶的酶学性质研究

采用超滤浓缩,乙醇沉淀,冷冻干燥和SephadexG-100层析等方法,获得香菇谷氨酰胺转氨酶粗酶及纯酶,对其酶学性质进行研究.结果表明:香菇中谷氨酰胺转氨酶粗酶的最适反应温度为40℃,温度稳定范围在50℃以下,最适PH为6.0,pH稳定范围在5.0～7.0,Na+、Ca2+等离子对该酶的活性影响甚微,为非Ca2+依赖性酶;纯酶的最适温度为40℃,以Nα-CBZ-Gin-Gly为底物时香菇谷氨酰胺转氨酶的Vmax为0.020mg/(mL·min),Km为1.520g/L.     

PURIFICATION AND CHARACTERIZATION OF A PROTEASE FROM THE PYLORIC CAECA OF MENHADEN (Brevoortia spp) AND MULLET (Mugil spp) FROM THE SOUTHWEST ATLANTIC REGION

A protease classified as trypsin was isolated and purified from the pyloric caeca of two species of fishes: Brevoortia spp (menhaden) and Mugil spp. (mullet). The characterization of both enzymes as trypsin was based on their molecular weight (24 kDa) determined by SDS-PAGE, their inhibition by some known trypsin inhibitors (PMSF, SBTI, TLCK and benzamidine), and their ability to hydrolyze the synthetic trypsin substrate N-α-Benzoyl-DL-arginine-p-nitroanilide (BAPA). Menhaden trypsin had maximum activity at pH 9.5 and was stable for 30 min between pH 6.0 and 10.0 at 0, 10 and 25C. On the other hand, mullet trypsin showed maximum activity at pH's from 7.8 to 9.0, and was stable over a wider pH range (7.0–10.0). The optimum temperature for menhaden and mullet trypsin was 63C and 60C respectively. Thermostability for menhaden trypsin was up to 50C, whereas mullet trypsin was stable up to 60C. The Km(BAPA) values for menhaden trypsin were conserved in the temperature range from 10 to 30C, while for mullet trypsin the Km(BAPA) was conserved between 10 and 25C.     

雪花粉及其后路高筋粉的品质特性研究

以永良4号小麦的前路粉（雪花粉）和后路粉（高筋粉）为原料，通过拉伸仪、粉质仪、发酵仪、面筋仪等分析手段对前路粉和后路粉的流变特性、发酵特性、凝胶特性及成品品质特性、蛋白质组成、多酚氧化酶的活性等进行了研究，发现雪花粉具有高的蛋白质量，适当的麦谷／麦胶、直链淀粉／支链淀粉比例，表现出对不同食品的适应性。以及低的多酚氧化酶活性表现出来的稳定性，高筋粉具有高的蛋白含量和质量，较差的发酵特性。通过搭配国产优质小麦面粉，加入适当的改良剂，可以生产出高品质的面包粉，搭配面筋含量高的普通面粉，或小麦面粉可以生产出优质的油条专用粉。     

PROPERTIES OF PHENOLOXIDASE ISOLATED FROM THE CEPHALOTHORAX OF KURUMA PRAWN (PENAEUS JAPONICUS)

Phenoloxidase (PO) from the kuruma prawn cephalothorax was partially purified and characterized. The enzyme showed maximal activity at pH 6.5 and 35C. It was stable in a wide pH range of 3–10 but unstable at a temperature greater than 50C. On the basis of activity staining with L‐β‐(3,4‐dihydroxylphenyl)alanine, the apparent molecular weight was estimated to be 160 kDa. Sodium dodecyl sulfate, methanol and trypsin showed no effect on PO activity, suggesting that the enzyme was isolated in the active form. Phenylthiourea, a copper‐chelating agent, and cysteine exhibited the inhibitory activity against PO in a concentration‐dependent manner. However, copper acetate addition had no influence on the activity.     

芒果叶中黄色素的提取及其稳定性研究

以芒果叶为原料,采用正交实验,建立了从芒果叶中提取黄色素的最佳工艺条件,并分析了该色素的稳定性.结果表明:提取溶剂为体积分数60%乙醇,料液比1:9(g:mL),提取温度60℃,提取时间1.5 h,提取次数为2次,浸提率为81.23%.芒果叶黄色素对酸、热稳定,Na 、Zn2 、氧化剂及食品添加剂(葡萄糖、柠檬酸、苯甲酸钠)对色素无影响,Fe3 、Cu2 和Vc对色素有一定影响.     

DIGESTION OF MYOFIBRILLAR AND SARCOPLASMIC FRACTIONS FROM MENHADEN MUSCLE BY THE ACTION OF MENHADEN (Brevoortia spp.) AND WHITE CROAKER (Micropogonias furnieri) TRYPSINS

Trypsins from the pyloric caeca of menhaden (Brevoortia spp.) and croaker (Micropogonias furnieri) were used for hydrolysis of both myofibrillar and sarcoplasmic fractions from menhaden muscle. Digestion of muscle proteins was carried out at 37C for 20 min with the pH maintained at either 3, 4, 5, 6, 7, 8 or 9; or for 60 min at the pH's of 5, 7, 8 or 9 at 30C and 37C. The hydrolytic action was evaluated based on the concentration of peptides solubilized. Solubility after digestion of myofibrillar, sarcoplasmic fraction and hemoglobin was optimum at basic pH. The sarcoplasmic fraction was solubilized more than the myofibrillar fraction by both fish trypsins. Similar results were obtained with both crude pyloric extracts and purified trypsins from menhaden and white croaker. Thus, the preparation of the crude pyloric caeca may be preferentially used, because of its low cost as well as the simplicity of the preparative procedure .     

PURIFICATION AND CHARACTERIZATION OF A SERINE PROTEINASE FROM THE TUNA PYLORIC CAECA

A 15.0 kDa serine proteinase with collagenase activity from pyloric caeca of tuna, Thunnus thynnus, was purified in four steps; acetone precipitation, gel filtration chromatography on a Sephadex G‐100, ion‐exchange chromatography on a DEAE‐Sephadex α‐50 and gel filtration chromatography on a Sephadex G‐75 column. The purification and yield were 30.5‐fold and 0.023%, respectively, as compared with those in the starting crude extract. The optimum pH and temperature for the purified collagenolytic enzyme were around pH 7.5 and 55C, respectively. The purified proteinase was strongly inhibited by metal ions (Hg²⁺ and Zn²⁺) and serine proteinase inhibitors (PMSF, TLCK and soybean trypsin inhibitor) suggesting it is a serine protease. The K_m and V_max of the purified enzyme for collagen type I were approximately 3.82 mM and 851.5 U, respectively.     

PURIFICATION AND CHARACTERIZATION OF TRYPSIN FROM PYLORIC CAECA OF BIGEYE SNAPPER (PRICANTHUS MACRACANTHUS)

Trypsin from the pyloric caeca of bigeye snapper was purified and characterized. Trypsin had an apparent molecular weight of 23.8 kDa when analyzed using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) and substrate‐gel electrophoresis. The trypsin fraction consisted of three isoforms as evidenced by the appearance of three different bands on native‐PAGE. Optimal activity was observed at 55C and pH range of 8–11. The activity of trypsin fraction was completely inhibited by soybean trypsin inhibitor and was partially inhibited by E‐64 and ethylenediaminetetraacetic acid. CaCl₂ partially protected the trypsin fraction from activity loss at 40C, while NaCl (0–20%) decreased the activity in a concentration‐dependent manner. The apparent Michaelis–Menten constant (K_m) and catalytic constant (k_cat) were 0.312 mM and 1.06 s, respectively when Nα‐Benzoyl‐dl ‐arginine ρ‐nitroanilide was used as a substrate. Trypsin from the pyloric caeca of bigeye snapper generally showed similar characteristics to other fish trypsins.     

COMPARATIVE STUDY OF STRUCTURAL CHARACTERISTICS AND THERMAL BEHAVIOR OF WHEY AND ISOLATE SOYBEAN PROTEINS

Electrophoretic profiles, sulfhydryl(SH)/disulfide (SS) groups content, and surface hydrophobicity (H₀) values of whey soybean proteins (WSP) and native soy isolates (NSI) were determined. WSP, composed mainly by Kunitz trypsin inhibitor (KTI), and lectin (L), has a H₀ value of 24.0 ± 1.0, which is 6.8 times lower than that of NSI ones, and SH/SS groups content in the same range of NSI. The thermal behavior of WSP and NSI was studied by differential scanning calorimetry (DSC). The WSP thermogram in water, similar to NSI, showed two main peaks (Tp values: 74.0 ± 0.5 C and 90.4 ± 0.8C) attributed to thermal denaturation of KTI and L, respectively. These endotherms are slightly affected by μ, whereas those of NSI are strongly affected (Tp of 7S and 11S peaks increase 17C and 20C respectively, by increasing NaCl concentration from 0 to 1M). WSP has low Ho values not noticeably affected by ionic strength changes, whereas NSI has higher Ho, that increase in saline media and favor intermolecular hydrophobic interactions. The consequent low tendency to protein aggregation by hydrophobic interactions in WSP would explain their lower thermal stability at high μ. Control proteins of both preparations (7S and 11S enriched fractions for NSI, and purified trypsin inhibitors, urease and lectin for WSP) were use to confirm these results.     

COMPARATIVE SENSORY ANALYSIS OF AQUACULTURED AND WILD YELLOW PERCH (PERCA FLAVESCENS) FILLETS

Tank-reared yellow perch in the round were bluish-green, and were coated with more slime than typical yellowish-green wild perch. Cooked fillets from aquacultured perch were whiter than those from wild fish, but were equal to wild fish in firmness and overall preference. Off-flavors were absent in aquacultured perch except in an instance when influenced by earthy flavors from microorganisms contaminating the recirculating water system.     

PARTIAL PURIFICATION AND CHARACTERIZATION OF A THERMOSTABLE TRYPSIN FROM PYLORIC CAECA OF TAMBAQUI (COLOSSOMA MACROPOMUM)

A 38.5 kDa alkaline protease from pyloric caeca of tambaqui (Colossoma macropomumj, a tropical freshwater fish, was partially purified in three steps: thermal treatment (45Cfor 30 min), salting‐out (ammonium sulfate at 40–80% of saturation) and gel filtration (Sephadex G‐75), The purification and yield were 51.2‐fold and 40%, respectively. The effects of pH, temperature, inhibitors, and substrates on proteolytic activities of partially purified enzyme were investigated. The optimum pH was 9.5, while the optimum temperature was 60C. This alkaline proteolytic activity remained unaltered after 30 min incubation at 55C. Active site inhibition provided additional evidence that this activity is attributed to a trypsin‐like enzyme.