Effect of Weaning Ages on Pancreatic and Intestinal Chymotrypsin Activity in Piglet

Twelve litters of new born piglets were divided randomly into groups I , I , M and F , and weaned at 17, 21, 28 and 35 days of age, respectively, to determine the effect of weaning age on pancreatic and intestinal chymotrypsin activity in piglets. The results showed that the relative and specific activity of pancreatic chymotrypsin increased significantly 12h after weaning compared with that of sucking piglets at the same age and then decreased and remained low for 2 - 3 weeks after weaning. Total pancreatic chymotrypsin activity had no change with the increase of pancreas weight during day 18 - 50 regardless of weaning (P > 0.05). Chymotrypsin activity of jejunum had the same change as that of pancreas during sulking stage. Jejunum chymotrypin activity decreased in the first week post-weaning in the piglets weaned before day 28, but had no change in groups weaned at 28 or 35 days. The earlier the weaning age, the longer the restoring time. Chymotrypsin activity in jejunum is more sensitive to the effects of weaning age than in duodenum and ileum.

SNP Analysis and Haplotype Identification in Chymotrypsin Inhibitor-2 (CI-2) Gene of Barley

Barley chymotrypsin inhibitor-2 （CI-2） was considered to be a promising candidate for enhancing the nutritional value of other cereals by increasing its concentration as it is rich in lysine than any other storage protein. Also, it was proposed that CI-2 might play an important role in the inhibition of proteolytic enzymes from pests or pathogens as CI-2 can strongly inhibit chymotrypsin and subtilisin. In this study, a total of 93 CI-2 gene sequences were isolated from wild and cultivated barley. 48 SNPs and 4 indels were detected across the entire sequences. The frequency of SNPs in the noncoding region （1 out of 9 bases） was slightly higher than that in the coding region （1 out of 10.7 bases）. In all, 33.3% of the candidate cSNPs resulted in amino acid changes. As a total, the 24 cSNPs resulted in 15 amino acid changes. Ten distinguishable haplotypes were detected, among which 3 haplotypes were shared in the most barley accessions, whereas the rest of the haplotypes appeared at a lower frequency. In addition, three haplotypes （haplotype 4, 8, and 9） were unique for single accessions. These results suggested that low diversity at the CI-2 locus was detected among the cultivated and wild barley.

Kinetic Resolution of DL-Phenylalanine Methyl Ester by α-Chymotrypsin Immobilized on Porous Silica Beads

Kinetic resolution of DL-phenylalanine methyl ester was carried out using drimobilized α -chymotrypsin (IC) as catalyst. The effects of temperatUre, pH, concentration of substrate and reactionvessels on the resolution were investigated. High quality L-phenylalanine was obtained in good yieldby an IC column.

<i>Fusarium lini</i>Potential for the Biotransformation of Norandrostenedione and Evaluation of Urease and Chymotrypsin Properties of the Transformed Products

Several androgenic steroids have been biotransformed by fungi into metabolites with numerous biological properties. Incubation of norandrostenedione (1) with Fusarium lini NRRL 2204 was carried out for the first time, yielding two new metabolites, 3,7β-dihydroxy-19-norandrost-1,3,5-trien-17-one (3) and 6α,10β,17β-trihydroxy-19-nor-4-androsten-3-one (4), along with three known compounds, 3-hydroxy-19-norandrost-1,3,5-trien-17-one (2), 10β, 17β-dihydroxy-19-nor-4-androsten-3-one (5) and 10β-hydroxy-19-nor-4- androsten-3,17-dione (6). Their structures were elucidated by extensive spectroscopic analyses, including 1D-, 2D-NMR, and HR-MS experiments. Substrate 1 and its derivatives 2-6 were evaluated in vitro for their urease and chymotrypsin inhibitory properties. Compounds 2 and 3 were found to have strong urease activity with IC50 = 23.7 ± 0.17 and 10.2 ± 0.28 μm, respectively, as compared to the standard drug thiourea (IC50 = 21.6 ± 0.12 μm). Compounds 4, 5 and 6 showed good chymotrypsin activity with IC50 values of 6.4 ± 0.19, 15.6 ± 0.46 and 18.4 ± 0.65 μm, respectively, as compared to standard chymostatin with IC50 = 5.7 ± 0.14 μm. These transformed metabolites may form the basis for the future development of new drugs against ulcer, inflammation, bacterial and viral diseases.

Comparison of the Contributions of Tetrahydrofurfuryl Alcohol and PEG to a-Chymotrypsin Renaturation with High Performance Hydrophobic Interaction Chromatography

The contributions of tetrahydrofurfuryl alcohol (THFA) and polyethylene glycol (PEG) to the renatured efficiency of a-chymotrypsin were investigated and compared with each other. The maximum increments of bioactivity recovery of a-Chy were found to be 25.1% for THFA, 10.4% for PEG, respectively. The experimental results indicated that the denaturant solution containing THFA contributed more to the renaturation of a-Chy in high performance hydrophobic interaction chromatography (HPHIC) than that containing PEG, when the concentration of THFA was 3.2%, the bioactivity recovery of a-Chy is the highest.

Adsorption of α-Chymotrypsin on Plant Biomass Charcoal

The adsorption of α-chymotrypsin onto plant biomass charcoal (PBC), which was prepared from plant biomass wastes such as bagasse and dumped adzuki beans by pyrolysis, has been examined. The PBC was characterized by SEM, specific surface area, and pore size distribution. The adsorption isotherms were successfully correlated by the Freundlich equation. The amount of α-chymotrypsin adsorbed on PBC was dramatically dependent upon the solution pH and temperature. Maximum adsorptions of α-chymotrypsin on adzuki bean charcoal and bagasse charcoal were observed at weak acidic and near neutral pH, respectively. The amount of α-chymotrypsin adsorbed on PBC decreased with an increase in the concentration of salts. Plots of the amount of α-chymotrypsin adsorbed on PBC versus temperature exhibited an optimum temperature.

Heat-Resistant Properties of <i>&#945</i>-Chymotrypsin Adsorbed onto Biomass Charcoal Powder

Biomass charcoal powder (BCP) was used as a carrier for immobilization of α-chymotrypsin through adsorption. BCP was derived from plant biomass wastes such as dumped bamboos by oxygen-free pyrolysis at low temperatures and grinding with a jet mill. The activity of adsorbed α-chymotrypsin was strongly dependent upon the kind of BCP. The thermal denaturation curve of adsorbed α-chymotrypsin was shifted to high temperature, compared to that of free one. When α-chymotrypsin adsorbed onto BCP of bamboos was incubated at 45°C, the half-life of adsorbed α-chymotrypsin was 2.6 times greater than that of free one. After incubation at 45°C, the remaining activity of adsorbed α-chymotrypsin markedly depended on the kind of BCP.

甲酸对丝素蛋白结晶结构转变的影响

结晶结构是丝素蛋白材料表征的重要内容,对丝素蛋白结晶结构的认识和调控是制备高性能材料的基础。甲酸作为一种可溶解蚕丝的优良溶剂,所制备的再生丝素蛋白材料具有优异的性能,其根源与甲酸对丝素蛋白具有促结晶的作用有关。文章对比分析了基于氯化钙-甲酸溶剂溶制再生丝素蛋白膜与基于三元溶剂溶制再生丝素蛋白膜的结晶结构差异,并利用α-糜蛋白酶从基于三元溶剂制备的丝素蛋白水溶液中分离出高结晶部分和低结晶部分,随后分别利用甲酸和乙醇对分离所得高结晶部分和低结晶部分进行处理,对比分析了乙醇和甲酸对其结晶结构转变的影响。结果表明,相对于乙醇处理,甲酸具有更显著的促丝素蛋白结晶的作用,更有利于丝素蛋白从无规卷曲或Silk I结晶态转变为SilkⅡ结晶态。

Chromosomal localization of silkworm (Bombyx mori) sericin gene 1 and chymotrypsin inhibitor 13 using fluorescence in situ hybridization

The chromosomal locations of two single-copy genes, Ser-1 and CI-13, in silkworm (Bombyx mori) were detected at the molecular cytogenetics level by fluorescence in situ hybridization in the study. The results showed that Ser-1 is located near the distal end of the 11th linkage group, relatively at the 12.5±1.4 position in pachytene; and that CI-13 has been mapped near the distal end of the 2nd linkage group, relatively at the 8.2±1.2 position in pachytene. Furthermore, their location model map-FISH map on silkworm chromosome was drawn. The FISH technique and its application to silkworm are also discussed in this paper.

Characterization and transcriptional analyses of cDNAs encoding three trypsin- and chymotrypsin-like proteinases in CrylAb-susceptible and CrylAb-resistant strains of sugarcane borer, Diatraea saccharalis

Diatraea saccharalis is a major corn borer pest. Midgut serine proteinases are essential for insect growth and development. Alteration of midgut proteinases is re- sponsible for Bt resistance development in some species. To clone midgut trypsin and chymotrypsin cDNAs and to test if the CrylAb resistance in D. saccharalis is associ- ated with changes in midgut proteinases, total midgut tryptic and chymotryptic activities, cDNA sequences, and gene expressions of three trypsin and three chymotrypsin genes were comparatively examined between Cry 1 Ab-susceptible （Cry 1 Ab- S S） and Cry 1 Ab-resistant （Cry 1 Ab-RR） strains. Full-length cDNAs encoding three trypsin- and three chymotrypsin- like proteinases were sequenced from CrylAb-SS and CrylAb-RR larvae. These cDNAs code for active forms of midgut serine proteinases with all fimctional motifs, includ- ing signal peptide, conserved His-Asp-Ser for the catalytic triad, three pairs of cysteines for disulfide bridge configurations, and conserved substrate specificity determination residues. In general, cDNA and putative protein sequences are highly similar between CrylAb-SS and CrylAb-RR strains, except for a few nucleotide and predicted amino acid substitutions, whose function need to be further clarified. Total trypsin and chymotrypsin activities were also similar between CrylAb-SS and CrylAb-RR strains. Transcriptional levels of the trypsin and chymotrypsin genes had numerical difference between Cry 1 Ab-SS and CrylAb-RR strains, but the difference was not statistically significant. Data suggest that the development of CrylAb resistance in D. saccharalis was not significantly as- sociated with these trypsins and chymotrypsins. Results clarified the role of six midgut proteinases and provided a foundation for continuing examination of potential involvement of other midgut proteinases in Bt resistance development and other important biochemical processes.

Synthesis of 2,3-dihydroquinazolin-4(1H)-ones catalyzed by a-chymotrypsin

We discovered that a-chymotrypsin has a promiscuous ability to catalyze the cyclocondensation of aromatic and aliphatic aldehydes with 2-aminobenzamides to afford the corresponding 2,3-dihydroquinazolin-4（1H）-ones successfully in high yields（90%–98%） under alcohol solvent. The catalytic activity of a-chymotrypsin was evaluated through investigating the temperature, the enzyme loading and the ratio of substrates in the enzyme-catalyzed reactions. The present method proves to be efficient and environmentally friendly in terms of short reaction time, high yield, green catalyst and the clean products obtained without further purification processes.

糜蛋白酶联合特布他林雾化治疗支气管哮喘的临床效果

目的探讨糜蛋白酶联合特布他林雾化治疗支气管哮喘的疗效及其对肺功能、炎性因子、免疫球蛋白水平的影响。方法将80例支气管哮喘患者随机分为对照组40例与观察组40例。2组均予以常规治疗,在此基础上对照组给予特布他林雾化治疗,观察组给予糜蛋白酶联合特布他林雾化治疗。比较2组临床疗效、临床症状消失时间及治疗前后肺功能指标[第1秒用力呼气容积(FEV1)、FEV1/用力肺活量(FVC)、呼出气一氧化氮(FENO)]、血清炎性因子水平[嗜酸细胞阳离子蛋白(ECP)、γ干扰素(IFN-γ)、白细胞介素-4(IL-4)、白细胞介素-1β(IL-1β)、超敏C反应蛋白(hs-CRP)]、免疫球蛋白水平[免疫球蛋白A(IgA)、免疫球蛋白M(IgM)、免疫球蛋白G(IgG)、免疫球蛋白E(IgE)]的变化;观察2组治疗期间不良反应发生情况。结果观察组治疗总有效率较对照组显著增加(P<0.05),症状消失时间较对照组显著缩短(P<0.05)。2组治疗后FENO值及血清ECP、IL-4、IL-1β、hs-CRP、IgE水平均较治疗前显著降低,FEV1、FEV1/FVC值及血清IFN-γ、IgA、IgM、IgG水平均较治疗前显著升高,且观察组治疗后上述指标较对照组治疗后改善更为显著,差异均有统计学意义(P<0.05)。2组治疗期间不良反应发生率比较差异无统计学意义(P>0.05)。结论糜蛋白酶联合特布他林雾化治疗支气管哮喘疗效确切,可在短时间内改善临床症状,促进肺功能恢复,缓解炎性反应,改善免疫功能。

Chymotrypsin B is not just a digest enzyme but widely expressed in rat tissues

The functions of lysosomal proteolytic enzymes were once believed to be limited to intracellular protein degradation occured within the lysosome. Recent findings suggest

Effect of ligand structure of stationary phase of high performance hydrophobic interaction chromatography on renaturation efficiency of GuHCl-denaturedα-chymotrypsin

The renaturation of the denatured α-chymotrypsin (α-Chy) with 1.7 mol · L-1 guanidine hydrochloride (GuHCI) by three kinds of stationary phase of high performance hydrophobic interaction chromatography (STHIC) with a comparable hydrophobicity but different ligand structures was investigated. The obtained result indicates that the ligand structures of the three STHIC contribute to the renaturation efficiency of α-Chy in the order of the end ligands PEG-600＜ phenyl group ＜ tetrahydrofurfuryl alcohol (THFA).

注射用糜蛋白酶在尿道成形术后患儿伤口换药中的应用

目的探讨注射用糜蛋白酶在尿道成形术后患儿伤口换药中的应用效果。方法选取2020年6月至2022年3月于江西省儿童医院泌尿外科接受尿道成形手术的88例尿道下裂患儿,将其随机分为试验组和对照组,各44例。试验组和对照组伤口换药前分别用16 mL的注射用糜蛋白酶稀释液(稀释于生理盐水中)和生理盐水浸润阴茎包扎敷料,敷料完全软化后予以换药。比较2组敷料完全软化所需时间、敷料与伤口粘连发生率、伤口出血发生率及换药时的疼痛程度[根据患儿年龄采用儿童疼痛行为量表(FLACC)、Wong-Baker面部表情疼痛评定量表(FPS)或视觉模拟量表(VAS)进行评估]。结果试验组患儿敷料完全软化时间较对照组明显缩短(P=0.006),且敷料与伤口粘连发生率、伤口出血发生率及换药时疼痛评分均低于对照组(均P<0.05)。结论注射用糜蛋白酶应用于尿道成形术后患儿伤口换药可加速血块溶解,加快敷料软化,缩短敷料浸润时间,减少敷料与伤口粘连及伤口出血的发生率,减轻疼痛。

前蛋白转化酶枯草溶菌素9通过肝源性巨噬细胞移动抑制因子调控动脉粥样硬化炎症反应的研究

目的利用人细胞株,系统研究PCSK9是否通过肝源性MIF调控单核/巨噬细胞炎症反应,进而为动脉粥样硬化(Atherosclerosis)等炎症性疾病的临床防控提供新的思路和依据。方法利用小干扰RNA(Small interfering RNA,SiRNA)介导的MIF基因转染THLE-3细胞,将实验分为MIF低表达组(Si-MIF-THLE-3)和MIF正常表达组(Con-THLE-3),通过实时荧光定量PCR(quantitative real-time PCR,RT-qPCR)检测肝细胞中PCSK9、MIF mRNA表达。在Si-MIF-THLE-3组和Con-THLE-3组中分别给与PCSK9重组蛋白进行干预,通过Western Blot检测肝细胞中PCSK9、MIF蛋白表达,并用ELISA测定培养液中PCSK9及MIF的含量,并收集上述肝细胞的培养液,-80℃冻存。利用SiRNA介导的MIF基因转染THP-1细胞,将实验分为MIF低表达组(Si-MIF-THP-1)和MIF正常表达组(Con-THP-1),通过Western Blot检测MIF的蛋白表达。将收集的肝细胞培养液分别加入Si-MIF-THP-1和Con-THP-1组共培养48 h,其中PCSK9干预组另设TLR4特异性抑制剂TAK-242(1μM)(P+T)。通过ELISA检测炎症因子表达;Western Blot检测THP-1细胞株中TLR4—NF-κB信号通路的表达。结果①Si-MIF-THLE-3组较Con-THLE-3组的MIF mRNA表达显著降低,而PCSK9的mRNA表达升高(P均<0.05)。②PCSK9干预后肝细胞MIF蛋白的表达以及分泌至培养液中的含量升高,而PCSK9蛋白的表达以及培养液中的含量不变(P均<0.05)。③Si-MIF-THP-1组较Con-THP-1组的MIF蛋白表达显著降低(P均<0.05)。④Si-MIF-THP-1组较Con-THP-1组炎症因子的表达低,PCSK9干预组较正常组和P+T组的炎症因子表达升高;PCSK9干预组较正常组和P+T组的TLR4蛋白表达升高;Si-MIF-THP-1组较Con-THP-1组的IKBα蛋白表达高,PCSK9干预组较正常组和P+T组的IKBα蛋白表达低;PCSK9干预组较正常组和P+T组的P65/P50蛋白表达高(P均<0.05)。结论PCSK9可诱导肝细胞MIF的合成与分泌,肝细胞分泌的MIF可与巨噬细胞表面受体TLR4结合,激活下游NF-κB炎症信号通路,促进TNF-α、白介-6、白介-1β、单核细胞驱化因子-1等炎症因子的分泌,激发炎症效应。

胰凝乳蛋白酶SplB在枯草芽孢杆菌中的重组表达及应用

通过启动子优化、宿主筛选,构建了C端带His-tag的胰凝乳蛋白酶类蛋白酶SplB表达载体,成功实现了SplB在枯草芽孢杆菌中的重组表达,对纯化的重组SplB进行酶学性质研究,并实现了重组蛋白酶SplB在重组蛋白Prx标签切割中的应用。结果表明,以枯草芽孢杆菌ATCC6051Δ10为宿主,通过启动子P_(43)介导的重组蛋白酶SplB表达活力最高(10.24 U/mL)。通过亲和层析纯化了重组表达的SplB,并进行酶学性质研究,其最适温度为40℃,最适pH值为8.5,且具有良好的温度和pH值稳定性。在离子浓度较低情况下,Co^(2+)对重组SplB活力有促进作用,Mg^(2+)、K^(+)不影响酶活力;而Cu^(2+)、Zn^(2+)、Ni^(+)离子抑制SplB活力;十二烷基硫酸钠极大抑制重组SplB活力。将重组SplB应用于切割重组蛋白Prx的标签,结果表明,重组SplB具有WELQ肽段标签切割作用,并且SplB浓度越高,目标条带越浓。本研究为优化SplB的重组表达及在食品、医药领域的应用提供支持。

纤支镜肺泡灌洗联合糜蛋白酶治疗小儿重症肺炎的临床效果

目的探究纤支镜肺泡灌洗联合糜蛋白酶治疗小儿重症肺炎的临床效果。方法方便选取2017年3月—2020年5月福建医科大学附属泉州第一医院收治的136例重症肺炎小儿随机均分为参照组和研究组,各68例。参照组采用纤支镜肺泡灌洗治疗,研究组采用纤支镜肺泡灌洗联合糜蛋白酶进行治疗,对比两组治疗效果和机体血气相应指标。结果治疗后研究组总有效率为97.06%,高于参照组的86.76%,差异有统计学意义(χ^(2)=4.847,P=0.028);治疗后,两组患儿PaO_(2)、血液pH与治疗前相比均升高,且研究组PO_(2)、血液pH均高于参照组,差异有统计学意义(P<0.05);治疗后,两组患儿Lac、PaCO_(2)较治疗前均降低,且研究组Lac、PaCO_(2)均低于参照组,差异有统计学意义(P<0.05)。结论使用纤支镜肺泡灌洗联合糜蛋白酶治疗应用于小儿重症肺炎,可有效改善患儿症状,提高治疗有效性,可在临床上借鉴应用。

龙血竭散治疗乳腺脓肿的临床应用价值

目的探讨龙血竭散和糜蛋白酶联合应用于乳腺脓肿切开引流术换药中的临床价值.方法将100例行乳腺脓肿外科切开引流术患者随机分为对照组及实验组,每组50例.对照组患者脓腔愈合期间,在行外科处理时将雷夫诺尔纱条填塞于脓腔内,并充分引流.实验组患者换药时,应用糜蛋白酶溶液及龙血竭散进行处理,对比两组患者的治疗效果.结果两组患者治疗前后的脓腔直径变化、愈合时间、疼痛、治疗满意度比较差异具有统计学意义(P<0.05);两组患者治疗前后白细胞(WBC)、谷丙转氨酶及谷草转氨酶血液学检测变化差异无统计学意义(P>0.05).结论龙血竭散联合糜蛋白酶溶液的换药方式可加快乳腺脓肿外科切开引流术后切口的愈合时间,减轻患者疼痛,并有一定的抗炎作用,同时无明显肝功能改变,值得在临床工作中应用.

胰凝乳蛋白酶样蛋白酶体的活性作为前列腺癌生物标志物的潜在应用

目的:探讨胰凝乳蛋白酶样蛋白酶体(chymotrypsin-like proteasome,CLP)活性作为前列腺癌生物标志物的可行性。方法:检测在体和离体的蛋白酶体活性,同时检测核因子κB的抑制蛋白α(inhibitory protein α of nuclear factorκB,IκB-α)、Bcl-2相关X蛋白(BCL-2 associated X protein,Bax)和细胞周期蛋白依赖性激酶抑制因子(cyclin-dependent kinase inhibitor,p27)的表达。结果:与正常前列腺上皮细胞及对照小鼠前列腺组织相比,前列腺癌细胞和前列腺肿瘤异种移植物中蛋白酶体底物蛋白IκB-α、Bax和p27的表达水平降低,CLP的离体和在体活性分别升高70%和23%。结论:CLP的活性可能是一种潜在的前列腺癌生物标志物,可能适合前列腺特异性抗原(prostate-specific antigen,PSA)在前列腺癌诊断中的补充。