Bromelain对HL-60细胞的诱导分化作用

目的：为获得对ＨＬ－６０细胞具有高效低毒作用的天然诱导分化剂而进行的研究。方法：以菠萝酶Ｂｒｏｍｅｌａｉｎ为药物，对人前髓细胞性白血病细胞株ＨＬ－６０进行作用，通过不同浓度及不同作用时间的药物作用，统计ＨＬ－６０细胞的增殖率、生化变化及形态学的变化。结果：发现Ｂｒｏｍｅｌａｉｎ在高浓度下（８μｇ／ｍｌ）作用４天即可诱导ＨＬ－６０细胞发生显著的诱导分化，其ＮＢＴ阳性率及形态学指标均达到６０％以上，电镜下核异染色质浓聚，出现特异性颗粒。结论：Ｂｒｏｍｅｌａｉｎ对ＨＬ－６０细胞具有明显的诱导分化作用，但其作用机理仍需进一步研究。

Technology for Enzymolysis of Jellyfish Brain Protein by Bromelain

[Objective] This study aimed to investigate the optimal technology for en- zymolysis of jeUyfish brain protein by bromefain. [Method] Effects of enzymolysis temperature, the amount of added enzyme, solid/liquid ratio, enzymolysis time and pH on the enzymolysis of total protein were investigated through single factor ex- periments, and some technical parameters were optimized by orthogonal experi- ments. [Result] The results revealed that enzymolysis temperature of 55℃, the amount of added enzyme of 4 000 U/g, solid/liquid ratio of 1：4, the amount of added enzyme time for 5 h and pH 7 were the optimal combination of parameters for the enzymolysis; the average degree of hydrolysis obtained from the verification tests was 36.9%, indicating that the optimized results of the orthogonal experiment were credible. [Conclusion] This study provides the optimized conditions reliable theo- retical foundation and technical support for the exploitation and utilization of jellyfish resources as medicines or functional food in China.

Application of Bromelain Extract for Muscle Foods Tenderization

This study investigates the tenderizing effects of bromelain extract (BE) obtained from the top phase of an aqueous two-phase system comprising of 18% PEG 6000–17% MgSO4 in muscle foods. Uniform-sized chunks of beef, chicken, and squid were marinated with different concentrations of BE (0, 3, 7%, and 20%, w/w) for 1 h at room temperature. The physico-chemical and quality characteristics of the marinated samples were determined. Decreases in pH and moisture content were observed in the samples BE treated, but the TCA-soluble peptides content significantly increased (p < 0.05). Water holding capacity and cooking yields were observed in all of the tested samples, especially when the concentration of BE increased. In addition, a reduction of meat firmness and toughness were also observed in all of the samples when compared to the control (p < 0.05). Electrophoretic patterns also revealed extensive proteolysis and a reduction in number and intensity of the protein bands in all of the treated samples. At the microstructural level, tissue fibers were broken, cell membranes were more strongly degraded, the connections between the sarcolemma and the myofibrils were disappeared, and the generation of numerous gaps could be clearly observed when 20% (w/w) BE was added. The results show that the bromelain extract derived from the two-phase extraction of a pineapple peel could be used as an effective meat tenderizer.

Optimisation of beef tenderisation treated with bromelain using response surface methodology (RSM)

The purpose of this study is to determine the optimum condition for the tenderization of beef by bromelain using Response Surface Methodology (RSM). Initially, bromelain powder was produced from pineapple crown of variety N36. Production of the bromelain powder involves several process steps such as extraction, purification, desalting and freeze drying. The cube size beef of round part was treated with bromelain at different pHs of beef, immersion temperatures, bromelain solution concentrations, and immersion times according to the experimental design which was recommended by RSM of MINITAB software version 15. Beef tenderness was then measured by Texture Analyser. The MINITAB software Version 15 was used to optimise the tenderisation of beef by bromelain. The determination coefficient R2 was 99.97% meaning that the experimental data were acceptable. It was found that beef could be optimize tenderised 89.907% at the optimum condition at pH of beef of 5.4, immersion temperature of60℃, bromelain solution concentration of 0.1682% and immersion time of 10 minutes. The verification value of beef ten-derisation at the feasible optimum condition which was determined by experiment was 89.571%. Since the difference between the veri- fication and predicted values was less than 5%, therefore, the optimum condition for the tenderisation of beef predicted by MINITAB software Version 15 could be accepted.

Incorporation of Bromelain into Dermatological Bases： Accelerated Stability Studies

This work aimed the incorporation of the enzyme bromelain from pineapple peel into dermatological bases for potential therapeutic application. The enzyme was incorporated into Lanette cream and lotion, as well as Carbopol gel and Chemyunion cream and lotion, at a concentration of 0.5% （w/w）. These bases were subjected to centrifugation test and accelerated stability test during 90 days at 25 ℃（with and without sunlight）, 37 ℃ and 4 ℃, in order to evaluate bromelain stability in dermatological bases. Total protein content and enzymatic activity were measured. The bases were stable as its organoleptic characteristics （appearance, color, smell and sensitivity to touch） only when kept at 4 ℃ with activity remaining 95.5%, 84,9%. 77.7%, 73.8% and 72.3%, alter 90 days of testing in Carbopol gel, Lanette and Chemyunion cream, and Lanette and Chemyunion lotion, respectively. Based on the results, it was possible to incorporate bromelain into dermatological bases, and the activity was well preserved when these bases were kept in refrigerator at 4 ℃.

Purification of Bromelain Enzyme from Curaua （Ananaserectifofius LB Smith） White Variety, by Aqueous Two-Phase System PEG 4000/Potassium Phosphate

The Curaua. （Ananaserectifolius LB Smith） is a species belonging to the family Bromeliaceae. Occurs in the states of Para, Amazonas, Amapa, Acre, Mato Grosso and Goia s, has two varieties, purple and white, and its fibers are used in automotive and textile industries due to its strength, softness and lightweight. Currently, only the fiber is used in industry, the rest is considered waste However, this residue contains compounds with important properties to be discovered and studied. An enzyme complex found in this residue is bromelain, a group of proteolysis＇ enzymes with application in several areas such as in the food, pharmaceutical and cosmetics industries. Today, its use is aiming the pharmaceutical industry, the production of ointments, gels, creams and lotions because they offer a wide range of therapeutic efficacies： antiedemas, anti-inflammatory, antithrombotic and fibrinolytic activities. This work studied the enzyme purification and recovery presented in the leaves of Curaua by means of an ATPS （aqueous two-phase system） PEG 4000/potassium phosphate. The protein content was measured by Bradford reagent and the enzymatic activity was measured by the Biuret reagent. Batch assays were performed aiming the enzyme extraction and recuperation, using the partition coefficient as indicator. It was used pH 7.0, 8.0 and 9.0, varying the proportional composition between the polymeric and the saline phases （tie-lines）. The white variety of Curaua. was used. The best purification factor was 4.53 and about 113.04 μg/mL of total protein measured by Bradford.

Kinetic studies on recombinant stem bromelain

Stem bromelain is a plant thiol protease with several industrial and therapeutic applications. This current work presents kinetic studies of recombinant bromelain (recBM) expressed in Escherichia coli BL21-AI on foursynthetic substrates, N-α-carbobenzoxy-L-alanyl-p-nitrophenylester (ZANPE), N-α-carbobenzoxy-L-arginyl-L-ar-ginine-p-nitroanilide (ZAANA), N-α-carbobenzo-xy-L-phenylalanyl-L-valyl-L-arginine-p-nitroanili-de (ZPVANA) and L-pyroglutamyl-L-phenylalanyl-L-leucine-p-nitroanilide (PFLNA). Hydrolytic activities of recBM at various pH and temperature conditions were compared to that of commercial bromelain (cBM). Both enzymes demonstrated high activities at 45o C and pH 5 - 8 for recBM and pH 6 - 8 for cBM. recBM showed marginally lower Kmand slightly higher kcat/Kmfor ZAANA, ZANPE and ZPVANA in comparison to cBM.trans Epoxysuccinyl-L-leucylamido {4- guanidino}butane (E-64) severely affected recBM and cBM hydrolysis of the synthetic substrates by competitive inhibition with Kivalues of 3.6 - 5.1 μM and 5.5 - 6.9 μM for recBM and cBM, respectively. The evaluated properties of recBM including temperature and pH optima, substrate specificity and sensitivity to inhibitors or activators, satisfy the requisites required for food industries.

Purification Processes and Market Potential of Bromelain in Brazil

Bromelain is the denomination given to the group of endoproteases obtained from members of Bromeliacea family. These enzymes have a wide range of proven applications and have been an object of study for worldwide researchers for decades. Over the years, several different downstream processes were studied in order to determine which technique would be worthwhile to be implemented in Brazil and provide the national market with such product. The objective of the present study is to relate the main studies in Brazil that has proven that bromelain purification can be cost-effective, in addition to the well-known benefits owned by such enzymes, and highlight the applications that create their market potential in the Brazilian market.

基于菠萝蛋白酶的羊毛生物酶法防毡缩整理

针对微生物蛋白酶在羊毛酶法防毡缩整理过程中易造成羊毛纤维损伤的问题,本文采用植物蛋白酶(菠萝蛋白酶)与预处理结合的工艺对羊毛织物进行防毡缩整理。结果表明,菠萝蛋白酶在预处理的基础上能够有效改善羊毛织物的防毡缩性能,整理后的织物毡缩率为5.29%,强力损失率为4.10%。通过扫描电镜、亚甲基蓝着色等表征方法综合判断,菠萝蛋白酶作用于羊毛鳞片层,进而实现在降低羊毛织物毡缩率的同时将强力损失率控制在一个较小的水平。

低共熔溶剂对蛋白酶法亲水改性锦纶纤维的影响

针对锦纶织物亲水性较差的问题,采用商用菠萝蛋白酶对其表面进行改性。通过单因素变量实验对反应残液进行紫外光吸光度测试以及静态接触角测试,确定菠萝蛋白酶处理锦纶的优化工艺条件,即在蛋白酶质量浓度32 mg/mL,pH 6.0,温度50℃,反应时间18 h条件下,可实现织物表面的亲水化转变。在优化工艺的基础上使用体积分数为40%CHCL-2EG低共熔溶剂缓冲液体系辅助激活菠萝蛋白酶,并对水解效果进行测试分析,发现CHCL-2EG对锦纶纤维具有刻蚀作用,织物表面粗糙程度增加,织物表面亲水效果进一步增强,最终接触角降低36.7%。

Invulnerability of bromelain against oxidative degeneration and cholinergic deficits imposed by dichlorvos in mice brains

BACKGROUND： The present study elucidates the protective potential ofbromelain against dichlorvos intoxication in mice brains. Dichlorvos induces the oxidative stress by disproportionating the balance between free radicals generation and their scavenging in neurons which leads to neuronal degeneration. METHODS： In this study, mice were divided into four groups- group I （control）, group II （dichlorvos treated）, group III （bromelain treated） and group IV （exposed to both bromelain and dichlorvos both）. RESULTS： Dichlorvos treatment increased the levels ofthiobarbituric acid reactive substances （TBARS） and protein carbonyl content （PCC） which indicate the increased oxidative stress. Meanwhile, brain endogenous antioxidants and cholinesterases level was decreased after dichlorvos exposure. Levels of TBARS and PCC decreased whereas cholinesterases level was recorded to be elevated after bromelain exposure. CONCLUSION： Bromelain offered neuroprotection by decreasing oxidative stress and augmenting cholinesterases in mice brains. This study highlights the invulnerability of bromelain against oxidative and cholinergic deficits in mice brains.

Bromelain and Cardiovascular Risk Factors in Diabetes:An Exploratory Randomized,Placebo Controlled,Double Blind Clinical Trial

Objective： To assess whether the dietary supplement （bromelain） has the potential to reduce plasma fibrinogen and other cardiovascular disease （CVD） risk factors in patients with diabetes. Methods： This randomized placebo controlled, double blind, parallel design, efficacy study was carried out in China and investigated the effect of 12 weeks of bromelain （1,050 mg/day） on plasma fibrinogen, This randomized controlled trial （RCT） recruited 68 Chinese diabetic patients [32 males and 36 females; Han origin, mean age of 61.26 years （standard deviation （SD）, 12.62 years）] with at least one CVD risk factor. Patients were randomized into either bromelain or placebo group. While bromelain group received bromelain capsule, the placebo group received placebo capsule which consisted inert ingredient and has no treatment effect. Subjects were required to take 1,050 mg （3× 350 mg） of either bromelain or starch-tiffed placebo capsules, two to be taken （2 × 350 mg） after breakfast and another （350 mg） after dinner, daily for 12 weeks. Plasma fibrinogen, CVD risk factors and anthropometric indicators were determined at baseline and at 12 weeks. Results： The change in the flbrinogen level in the bromelain group at the end of the study showed a mean reduction of 0.13 g/L （standard deviation （SD） 0.86g/L） compared with the mean reduction of 0.36 g/L （SD 0.96 g/L） for the placebo group. However, there was no significant difference in the mean change in fibrinogen between the placebo and bromelain groups （mean difference=0.23g/L （SD 0.22 g/L）, P=0.291）. Similady, the difference in mean change in other CVD risk factors （blood lipids, blood pressure）, blood glucose, C-reactive protein and anthropometric measures between the bromelain and placebo groups was also not statistically significant. Statistical differences in fibrinogen between bromelain and placebo groups before the trial despite randomization may have influenced the results of this study. Conclusion： This RCT failed to show a beneficial effect in reducing fibrinogen or influencing other selected CVD risk factors but suggests other avenues for subsequent research on bromelain.

奶茶饮料中茶多酚含量测定的方法改进研究和应用

茶多酚含量是评价茶饮料质量的重要指标,国家标准《茶饮料》(GB/T 21733—2008)附录A采用乙醇沉淀-酒石酸亚铁比色法测定茶饮料中茶多酚的含量,该方法适用范围较小,特别是奶茶等使用了乳化剂、稳定剂的饮料产品,存在比色溶液浑浊的问题,直接影响检测结果的准确性。本研究通过改进样品预处理方法,使用菠萝蛋白酶酶解,解决了比色溶液浑浊的问题,提高了奶茶饮料中茶多酚含量测定的精密度和准确度。方法回收率为99.1%~102.0%,相对标准偏差RSD为0.23%~0.37%(n=6)。

前处理方式对冻干鸡肉制备及其特性的影响

为了改善冻干鸡肉的生产效率及特性,利用超声波、菠萝蛋白酶、菠萝蛋白酶-超声波联合进行前处理,探究其对冻干鸡肉的影响。结果表明,经超声波、联合处理样品的干燥速率高于未处理组,联合处理样品的总失水率显著高于其他组(P<0.05);联合处理样品的肌原纤维蛋白(myofibrillar protein,MP)溶解度以及电位绝对值显著高于其他组(P<0.05),其硬度、复水后嫩度、肌原纤维小片化指数(myofibril fragmentation index,MFI)、MP含量及平均粒径显著低于其他组(P<0.05);扫描电镜显示联合处理样品的肌原纤维断裂更明显;十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(sodium dodecyl sulfate-polyacrylamide gel electrophoresis,SDS-PAGE)显示联合处理能够降解肌球蛋白重链(myosin heavy chain,MHC)、肌动蛋白(actin)和原肌球蛋白(tropomyosin,TM),导致其条带变浅。联合处理有利于冻干鸡肉的制备生产以及感官、理化等特性的改善,能有效提高其品质。

An updated review of pineapple and its bioactive compounds in breast cancer

Women are more likely than men to develop cancer of the breast.Most breast cancer drugs are highly toxic,and treatment can cause side effects.It is imperative to find safe alternative medicines in the pursuit of a cure for breast cancer.An extract of pineapple contains cysteine proteases known as bromelains.In general,pineapples are regarded as safe foods.From the fruit,stem,and of pineapples,bromelain is produced by multiple endopeptidases.As well as reducing the growth of tumors locally,bromelain severely impaired the cytotoxicity of monocytes in the immune system in the fight against cancer.Specifically,we investigated pineapple’s possible mechanisms of action and its bioactive compounds in breast cancer.

菠萝蛋白酶的提取纯化工艺研究

为降低环境污染,提高菠萝蛋白酶的附加值以及延长菠萝的产业链,以菠萝废弃物为原料,采用盐析浓缩-透析提纯-乙醇有机溶剂沉淀相结合的方法提取菠萝蛋白酶。实验考察料液比、缓冲液温度、搅拌时间、缓冲液pH值、盐析浓度以及透析时间对菠萝蛋白酶酶活性和酶活回收率的影响。结果表明,菠萝蛋白酶的最佳工艺条件为料液比1.0∶1.5(m∶v)、缓冲液温度30℃、搅拌时间40 min、缓冲液pH值7、盐析浓度30%和透析时间28h,该条件下菠萝蛋白酶酶活回收率为68.45%。

超声波对菠萝果蛋白酶活性和光谱的影响

以菠萝果为材料,用盐析法制备菠萝果蛋白酶粗酶,进一步经Sephadex G-50和DEAE-纤维素柱层析纯化,其SDS·PAGE呈单一的电泳带,并获得了菠萝果蛋白酶结晶。一定参数的超声作用可以使酶活力提高,经光谱分析,超声处理后酶的吸收光谱不改变,而荧光发射峰由336nm红移到339nm,紫外差示光谱在一定的波长也呈现出明显的正峰和负峰。本文讨论了超声波影响菠萝果蛋白酶活性的机理。

化腐再生散促进糖尿病足肌腱坏死创面早期修复临床研究

[目的]观察化腐再生散对糖尿病足肌腱坏死创面早期的影响。[方法]71例糖尿病足患者,随机分为化腐再生散+生肌象皮膏组(A组)35例,单纯生肌象皮膏组(B组)36例,对其创面适时行蚕食清创后,分别采用不同药物治疗,观察14 d后坏死肌腱脱落情况以及治疗前后血常规中白细胞以及伤口分泌物中肿瘤坏死因子-α(TNF-α)等炎性指标的变化,同时对治疗前后伤口组织进行病理分析,综合分析药物对创面早期修复的影响。[结果]A组坏死肌腱治疗后较B组脱落明显;两组白细胞均呈下降趋势,但A组下降迅速,与B组比较差异有统计学意义(P<0.05);而两组分泌物TNF-α均呈上升趋势,A组较B组升高(P<0.05);病理学检测显示换药后A组新生血管生成较B组明显。[结论]应用化腐再生散结合生肌象皮膏换药治疗糖尿病足肌腱坏死创面,能够尽早促进坏死肌腱脱落,激活抗炎免疫机制,控制感染,促进新生肉芽组织生长,从而促进创面修复进程。

以波纹巴非蛤为原料制备海洋生物活性肽

目的:以波纹巴非蛤为原料,制备海洋生物活性肽,并确定最佳酶解条件。方法:选用木瓜蛋白酶和菠萝蛋白酶为水解酶,以水解度为考察指标,通过正交试验对酶解实验条件进行优化。结果:木瓜蛋白酶在50℃、酶解时间4h、加酶量4%、pH6.3、料水比1:3时水解度最高为40.83%,总氮回收率为89.85%,平均肽链长度为2.20;菠萝蛋白酶在温度55℃、酶解时间4h、加酶量12%、pH6.3、料水比1:3时水解度最高为34.61%,总氮回收率为89.58%,平均肽链长度为2.58。结论:在本实验条件下,木瓜蛋白酶对以波纹巴非蛤为原料制备海洋生物活性肽效果优于菠萝蛋白酶。

海藻酸钠包埋法制备固定化菠萝蛋白酶

以海藻酸钠为载体,包埋法固定菠萝蛋白酶,对固定化条件进行优化,同时探讨固定化菠萝蛋白酶的部分酶学性能。结果表明:固定化菠萝蛋白酶的质量受海藻酸钠质量分数、固定化酶量、固定化时间以及CaC l2质量分数的影响,其最佳固定化条件为:海藻酸钠质量分数1.0%,CaC l2质量分数3%,固定化酶液量与海藻酸钠体积之比1∶2,固定化时间60 m in,在此条件下,制备的固定化菠萝蛋白酶的比活力为211.8 U/g(湿质量载体),由此制得的固定化酶的最适pH为7.6,与游离酶相比,升高了0.8个pH单位,同时显示固定化菠萝蛋白酶能耐受较高的碱性环境,固定化酶最适温度与游离酶相同,均为50℃,固定化酶在较高温度范围内,仍能保持较高的相对活力。