海藻降血压肽的作用机制与制备

海藻是我国重要的海洋生物资源,因其含有丰富的活性物质成分而成为开发海洋药物研究的主要方向之一.来源于海藻中的降血压肽,安全无毒、无副作用,对预防和治疗高血压具有良好的效果.本文在介绍海藻中的降血压活性成分和降血压肽的作用机制的基础上,综述了海藻降血压肽的研究进展及其制备过程,并提出其开发应用前景.     

贻贝蛋白酶解降血压肽的降压活性及相对分子质量与氨基酸组成

以贻贝（Mytilus coruscus）为原料制取降血压肽,采用均匀设计试验探讨了不同酶解条件对其ACE( angiotensin converting enzyme) 抑制活性的影响,利用高效液相色谱分析了其相对分子质量分布,以氨基酸自动分析仪分析其氨基酸和游离氨基酸组成;并利用自发性高血压大鼠（SHR)饲喂试验检测其降血压活性。结果表明,对于P1酶来说, 酶量从25~175 IU• mL-1,酶解时间从2~7 h,随着酶解程度的提高,其ACE抑制率下降;其最高的ACE抑制率为88.08%,对应的最佳酶解条件为：酶量25 IU•mL-1,酶解时间2 h,pH 7.0,酶解温度60 ℃。此外发现降血压肽的相对分子质量在1.6 ku以下,其中10肽以下的短肽约占50.4%,证实了降血压肽主要以分子量较小的短肽组成。降血压肽游离氨基含量为8.8 g•（100g）-1,水解氨基酸总量为59.6 g•（100g）-1,疏水性氨基酸含量为24.86 g•（100g）-1,氨基酸组成比例平衡,必需氨基酸含量丰富,其中含量最高的氨基酸是谷氨酸,占8.57%。此外以3.0 g•kg-1体重的降血压肽样品剂量饲喂SHR,在饲喂后2～6 h降压效果显著,平均降低幅度为18～28 mmHg（P＝0.05）。研究表明以贻贝等水产蛋白为原料可开发具有高ACE抑制活性和显著降血压作用的降血压肽。     

祖国各地

国家海洋局二所开发出高活性降血压肽在国际科学基金资助及国家海洋局的支持下,国家海洋局第二研究所研究开发出一种高活性降血压肽。经研究,该所的科研人员发现,鲢鱼和贻贝中含有的肽可以治疗高血压症。他们以鲢鱼和贻贝作原料,采用分步复合酶酶解及活性修饰等先进工艺,从中提     

血管紧张素转换酶（ACE）的提取与活性验证

在从紫菜中提取制备降血压肽的研究中,为筛选能够抑制血管紧张素转换酶（ACE）活性的功能肽,需要大量ACE进行体外检测。但ACE成本昂贵,本文利用硫酸铵分级盐析法提取猪肺组织中的ACE,测其酶活力为15．45×10^-3U／mg,比活力为33．80×10^-3U／mg。通过卡托普利来验证其抑制效果,抑制率达49．15％。采用本实验室初步制得的紫菜降血压肽对自制ACE和商品ACE的活性进行抑制,测定IC50发现该抑制剂对2种ACE的抑制效果差异不大,且活性较为稳定,因此所提取的ACE粗酶液可直接用于降血压肽的体外检测实验。     

国外简讯

从大马哈鱼中提取降血压活性肽日本北海道食品加工中心研究成功从大马哈鱼头部提取降血压的保健药品与食品。提取方法是将大马哈鱼头洗净后捣碎、加蛋白酶磨成糜浆状，使蛋白质转化成活性肽，其分子量达３００。此种活性肽具有生理活性，能抑制血压上升因子—高血压蛋白Ⅰ...     

国家海洋局二所开发出高活性降血压肽

在国际科学基金资助及国家海洋局的支持下，国家海洋局第二研究所研究开发出一种高活性降血压肽。经研究，该所的科研人员发现，鲢鱼和贻贝中含有的肽可以治疗高血压症。他们以鲢鱼和贻贝作原料，采用分步复合酶酶解及活性修饰等先进工艺，从中提取出降血压肽，经冷冻干燥，制剂成型。经过对患有高血压症的老鼠进行试验，证明该药降压效果显著。对病鼠的给药剂量为3g／kg体重，6h后，老鼠的血压平均下降20～25mmHg，最大降幅达30mmHg。经安全性检测，证明此药安全、无毒副作用。     

紫菜降血压肽酶法制备工艺的优化

摘要：降血压肽又称为血管紧张素转化酶抑制肽,是具有降血压活性的生物活性肽,对高血压患者有显著疗效。为探讨从紫菜中制备降血压肽的酶法工艺,本实验选用AS．1398中性蛋白酶、胰蛋白酶、复合酶、木瓜蛋白酶、碱性蛋白酶、风味蛋白酶等6种蛋白酶分别对紫菜进行酶解,通过对其酶解液的ACE抑制活性和水解度的比较,发现采用AS．1398中性蛋白酶酶解的产物ACE抑制活性最高。对AS．1398中性蛋白酶的酶解工艺进一步优化,确定其最佳酶解工艺为：温度50℃、pH值7．5、加酶量E／S10000U·g^-1。在此条件下,酶解4h和10h的酶解产物抑制活性最高均达70％以上,并发现紫菜酶解液的ACE抑制率与水解度的增长不成显著的线性关系。     

源于海洋贝类蛋白的ACE抑制肽研究现状

ACE抑制肽在血压调节方面起着重要作用,而海洋贝类含有丰富的蛋白质,并具有开发ACE抑制肽的潜能。本文主要简述了源于海洋贝类蛋白源的ACE抑制肽的制备、分离纯化、活性评价及构效关系等方面,以期为海洋贝类蛋白的开发利用提供参考。     

金枪鱼肽可快速降血压

日本静冈县静冈市研究人员确认，金枪鱼可以降血压。研究人员从金枪鱼中提取精炼的金枪鱼肽具有快速降血压的功效。     

鱼降压肽的酶法制备工艺及其理化性质研究

采用高压液相色谱测定降血压肽的血管紧张素转化酶抑制活性,对碱性蛋白酶水解草鱼蛋白制备鱼降压肽进行研究。结果表明,碱性蛋白酶水解草鱼蛋白制备鱼降压肽的较佳工艺条件为：pH9.0、温度50℃、酶与底物比48AU.kg-1、水解度34.52%。鱼降压肽成分分析结果显示,鱼降压肽中可溶性氮含量为81.26%,肽含量为72.81%,脂肪为0.12%,水分为3.54%,灰分为9.47%。鱼降压肽的血管紧张素转化酶抑制活性为70.35%;体积排阻色谱测定鱼降压肽的相对分子质量在124～10581之间,主要集中在124～1062之间;溶解度的测定结果表明,鱼降压肽在pH3.0～11.0时溶解度为96.0%左右,可广泛应用于食品中。 关键词：草鱼; 降压肽;碱性蛋白酶;高压液相色谱;相对分子质量;体积排阻色谱     

Rare serotype occurrence and PFGE genotypic diversity of Streptococcus agalactiae isolated from tilapia in China

Previously, we reported 10 PEGE types of 85 tilapia Streptococcus agalactiae(GBS), which shifted from Streptococcus iniae in China, by using PEGE method. Presently, larger and more representative tilapia GBS were isolated, for the ?rst time in China, to characterize their serotypes and genetic diversities more precisely than had done before. 168 GBS strains were distributed in ?ve provinces of China, in which Guangdong, Guangxi and Hainan were the major ones, holding36.9%(62/168), 37.5%(63/168) and 19.6%(33/168), respectively. Serotypes, Ia, Ib and III, were observed in these strains and the most predominant one was Ia(95.2%), which mainly distributed in Guangdong, Guangxi and Hainan. Ia initially occurred in 2009, it shoot up to 32.1% in 2010,but decreased to 16.1% in 2011 before went up to 45.2% in 2012. Ib sporadically occurred during2007–2011, III onlyoccurred in 2012. 14 different PFGE types, including 4 new types(N, O,P and Q), were observed, in which B, D, F and G were the predominant types, holding 83.9%(141/168) of the total GBS strains. Ia corresponded to 11 PFGE types(A–H, N–P), in which type D predominated(51%). Ib represented 3 genotypes(I, J and Q) and III harbored only 2genotypes(N and F). Type N and Fsynchronously presented in Ia and III. In summary, the genetic diversity of tilapia GBS varied by serotypes and changed with geographical locations and years.Although Iastillpredominated, new rareserotypeIII alreadyoccurred in China.     

Growth hormone (GH) and reproduction: a review

Interaction between growth and reproduction occurs in many vertebrates and is particularly obvious at certain stages of the life cycle in fish. Endocrine interactions between the gonadotropic axis and the somatotropic axis are described, the potential role of GH being emphasised. A comparative analysis of these phenomena in mammals, amphibians and fish, suggests a specific role of GH in the physiology of puberty, gametogenesis and fertility. It also shows the original contribution made by studies on the fish model in this field of investigations.     

Purification and partial characterization of GtHs (cfLH and cfFSH) from Indian walking catfish (Clarias batrachus) (L.) and development of a homologous ELISA for cfLH

Two gonadotropins (GtH; Qa and Qb) were purified by gel filtration and ion exchange chromatography from the pituitaries of Indian walking catfish (Clarias batrachus). The presence of GtH during purification was assessed by in vitro oocyte maturation and in vivo steroidogenic activity, and their identities were determined by elution profiles, molecular weight, biological activities and yield. The molecular weights of Qa and Qb were 37 and 42 kDa, respectively, and composed of distinct subunits (Qa: 20 and 14 kDa and Qb: 26 and 18 kDa). Polyclonal antibodies raised against Qa immunostained Qa, Qb and pituitary GtH cells. A competitive Qa‐ELISA was developed whose sensitivity was 6.25 ng mL^?1 (1.25 ng well^?1) with intra‐ (3.5%) and inter‐ (12.4%) assay coefficients of variation. Displacement curves parallel to the standard were obtained with plasma and pituitary extracts of catfish, Qb and carp GtHII. The assay was validated by measuring the plasma Qa levels after LHRH treatment and in relation to ovarian growth in the female catfish during different reproductive phases. Based on the results, Qa and Qb corresponded to fish LH and FSH respectively. The findings will increase the knowledge of the mechanisms controlling fish reproduction and identification of sensitive phases in fish in captivity for hormonal manipulation.     

Controlled infection of Poecilia reticulata Peters (guppy) with Tetrahymena by immersion and intraperitoneal injection

Tetrahymena is a protozoan parasite, which infects guppy, Poecilia reticulata Peters, and causes substantial economical losses in commercial farms worldwide. Studies of guppy infected by Tetrahymena require standardized infection protocols. The LD50 for Tetrahymena infection of guppies by intraperitoneal (IP) injection was calibrated, and the level obtained was 946 parasites per fish. Guppy infection with Tetrahymena by immersion, imitating the natural route of infection via the integument, was studied under normal or stress conditions. Exposure to cold and netting (CNI) and to cold only (CI) followed by immersion exposure to 10 000 Tetrahymena per mL resulted in 22.5% and 19.2% mortality, respectively, as compared to 14.2% and 10% in groups that were netted only (NI) or non‐stressed (I). Histopathology revealed that immersion infection resulted in a systemic infection. Lysozyme levels, measured 3 weeks after infection, were significantly higher in the CNI group (288 μg per mg protein) compared with CI‐, NI‐ and I‐treated groups (94.5, 64 and 62.3 μg mg^?1, respectively). There was no evident parasite immobilization activity in body homogenates, suggesting no development of acquired immunity. Re‐infection by IP injection revealed no increase in protection in any of the treatment groups, mortality range of 56.3–75%, higher than in the non‐exposed control (40.6% mortality).     

Imported ornamental fish are colonized with antibiotic‐resistant bacteria

There has been growing concern about the overuse of antibiotics in the ornamental fish industry and its possible effect on the increasing drug resistance in both commensal and pathogenic organisms in these fish. The aim of this study was to carry out an assessment of the diversity of bacteria, including pathogens, in ornamental fish species imported into North America and to assess their antibiotic resistance. Kidney samples were collected from 32 freshwater ornamental fish of various species, which arrived to an importing facility in Portland, Oregon from Colombia, Singapore and Florida. Sixty‐four unique bacterial colonies were isolated and identified by PCR using bacterial 16S primers and DNA sequencing. Multiple isolates were identified as bacteria with potential to cause disease in both fish and humans. The antibiotic resistance profile of each isolate was performed for nine different antibiotics. Among them, cefotaxime (16% resistance among isolates) was the antibiotic associated with more activity, while the least active was tetracycline (77% resistant). Knowing information about the diversity of bacteria in imported ornamental fish, as well as the resistance profiles for the bacteria will be useful in more effectively treating clinical infected fish, and also potential zoonoses in the future.     

Effect of iodophor disinfection of non‐hardened Salmo trutta eggs on their bacterial and fungus load

This study investigated the efficiency of iodophor disinfection (135 ppm active iodine for 15–30 min) of non‐hardened Salmo trutta eggs against different groups of bacteria and against fungus. Egg samples were taken from non‐disinfected and from disinfected eggs, microorganisms were cultured on specific nutrient media and their mass was measured by turbidimetric methods. Bacteria and fungus mass of non‐hardened eggs could be reduced but not eliminated by iodophor disinfection with 135 ppm active iodine for 15 min. The extent of reduction was 47–65% (Experiment 1). The efficiency of disinfection increased with disinfection time as the reduction in bacteria and fungus mass was 40–55% after 15 min and 58–74% after 30 min (Experiment 2). Disinfection efficiency of iodophor solution diluted in water (reduction 49–57%) and of iodophor solution diluted in sodium chloride solution iso‐osmolar to the oocytes (reduction 52–61%) was similar (Experiment 3). The reduction in bacteria and fungus mass was persistent as it was 39–72% lower in embryos deriving from disinfected eggs than in embryos deriving from non‐disinfected ones (Experiment 4). In conclusion, the tested disinfection method is inadequate to eliminate pathogens completely but it could positively influence immune defence of eggs and embryos.     

Nitrogenous waste excretion and accumulation of urea and ammonia inChalcalburnus tarichi (Cyprinidae), endemic to the extremely alkaline Lake Van (Eastern Turkey)

The endemic, anadromous cyprinidChalcalburnus tarichi is the only fish species known to occur in alkaline Lake Van (Eastern Anatolia, Turkey). EightC. tarichi were maintained individually in Lake Van water (17 – 19°C; pH 9.8; 153 mEq·I^–1 total alkalinity; 22 total salinity) and tank water samples analyzed for 24 h in 2 to 4 h intervals. At zero time, < 1µM ammonia was present and urea was undetectable in the tank water; at 24 h, total ammonia and urea made up 114±32 and 35±25µM, respectively. Over the experimental period, ammonia-N and urea-N excretion averaged 1041±494 and 607±169moles·kg^–1 fish·h^–1, respectively. The extent of urea excretion was highly variable between specimens. Uric acid excretion was not detectable.Urea was present at high concentrations in all tissues and plasma (25 – 35moles·g^–1·ml^–1) of freshly caughtC. tarichi; total ammonia content of the tissues was by a factor of 1.9 (liver) to 3.0 (brain) lower. High arginase activity (2.4±0.2 U·min^–1·g^–1) was detected in the liver ofC. tarichi but ornithine carbamoylphosphate transferase, a key enzyme of the ornithine-urea-cycle, was absent. Ureagenesis is likely through degradation of arginine and/or uricolysis. High glutamine synthetase activity (11±0.6 U·min^–1·g^–1) and low ammonia content in brain suggest that, like other teleosts,C. tarichi has an efficient ammonia detoxification in the brain, but in no other tissue.Nitrogenous waste excretion at alkaline pH is discussed. The ability ofC. tarichi to excrete high levels of ammonia at extremely alkaline pH is unique among teleosts studied so far. The mechanism of ammonia excretion under Lake Van conditions remains to be elucidated.     

Influence of the antibacterial herbs, Solanum trilobatum, Andrographis paniculata and Psoralea corylifolia on the survival, growth and bacterial load of Penaeus monodon post larvae

The indiscriminate use of antibiotics and chemicals in shrimp hatcheries has led to biomagnification and that in turn could lead to rejection of a whole consignment. The application of the bioencapsulation technique as a tool for curative treatment in shrimp larvae was investigated. Herbs having antibacterial properties such as Solanum trilobatum, Andrographis paniculata and Psoralea corylifolia (methanolic extracts) were bioencapsulated in Artemia and fed to Penaeus monodon post larvae PL 1–25. The post larvae were reared in a medium inoculated with pathogenic bacteria such as Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella typhi and Vibrio sp. Post larvae reared in the non-inoculated water and fed with non-enriched Artemia exhibited 90% survival, highest specific growth rate (12.43%) and reduced bacterial load. P. monodon reared in the bacterial inoculated water and fed with the non-enriched Artemia exhibited the lowest survival (10–30%), specific growth rate (8.42–9.1%) and increased bacterial load (2.86 × 10³ to 3.76 × 10⁵ cfu/g). The methanolic extracts of the herbs helped to increase survival and specific growth rate and reduced bacterial load in the P. monodon culture system. Among the three herbal extracts, P. corylifolia enriched Artemia fed post larvae showed the tendency to higher survival (>50%), growth rate (11.5 averaged) and low bacterial load (1.12 × 10⁵ cfu/g). This revised version was published online in August 2006 with corrections to the Cover Date.