THE ROLE OF ACTIVE AND INACTIVATED PAPAIN IN BEER CHILLPROOFING

The mechanism of chillproofing of beer by papain has been studied in a series of three experiments. Changes in the molecular size of proteins and polypeptides have been monitored by chromatographic profiles on Sephadex G-25 columns. Simultaneously the production of free amino acids and changes in the colloidal stability of the beer were determined. The addition of active papain to beer affected all three parameters. By contrast when inactivated papain was added to beer neither the chromatographic profile nor the concentration of free amino acids were altered but an increase in colloidal stability occurred. The removal of active papain from beer did not affect its colloidal stability but the removal of inactivated papain from beer stabilised with this agent eliminated the chillproofing effect.     

木瓜蛋白酶在亲和双水相系统中的分配行为及机制研究进展

本文查阅相关文献, 对木瓜蛋白酶传统提取方法(超滤法、盐析法、有机溶剂法、亲和层析法)及一些新兴的提取方法如双水相萃取法、亲和双水相法作了简要综述。利用金属螯合亲和双水相分配系统, 能有效获得高纯度木瓜蛋白酶, 但木瓜蛋白酶与亲和成相剂的相互作用及其分配行为与机制尚不清楚。研究木瓜蛋白酶与不同金属离子亲和成相剂之间的相互作用和吸附动力学模型;对酶的亲和结合物进行物理表征和分子模拟。阐明木瓜蛋白酶与不同金属离子亲和成相剂之间的亲和作用原理。将双水相萃取技术和亲和分配技术相融合, 提出基于木瓜蛋白酶的金属螯合亲和双水相分配技术, 该技术有望大力推动木瓜蛋白酶工业化高效制备的发展。     

Determination of papain in raw meat by immunoassay

An antibody raised to papain, a meat tenderising proteolytic enzyme obtained from papaya (Carica papaya), has been used in the development of an enzyme-linked immunoabsorbent assay (ELISA) for the determination of papain in raw meat. Quantitative determinations of papain up to 4 mg/kg of raw meat have been obtained using standard extracts prepared by the exogeneous addition of papain to raw beef. A sample of commercially treated 'tenderised beef' was shown to contain papain at the level of 0·40 mg/kg. In collaboration with a Public Analyst, a papain immunoassay kit has been used to assay 50 samples of beef bought from retail outlets, with a view to monitoring the use of this tenderiser by the meat industry.     

血红蛋白酶法脱色技术研究

为充分改善血红蛋白的感官质量并扩大其应用范围,本实验以猪血为原料,以猪血红蛋白为研究对象,采用6种食品级商业蛋白酶Alcalase 2.4L,胰蛋白酶,Flavourzyme,AS1398中性蛋白酶,木瓜蛋白酶及胃蛋白酶在各自最适反应条件下分别酶解猪血红蛋白480min,以产物色泽为评价指标,筛选出木瓜蛋白酶为最佳水解用酶。其酶解液接近无色,透明、清澈,感官质量得到了很大提升。其酶解条件为底物浓度5%,酶与底物比100U/g,温度37℃,pH值6.5。各蛋白酶水解液的色泽由浅至深依次为:木瓜蛋白酶,Flavourzyme,AS1398中性蛋白酶,胰蛋白酶,胃蛋白酶和Alcalase 2.4L。     

壳聚糖固定化木瓜蛋白酶提取牛蒡多糖的研究

以壳聚糖为载体、戊二醛为交联剂固定化木瓜蛋白酶,从牛蒡中提取多糖,考察固定化工艺的选择、固定化酶提取牛蒡多糖的优化条件及固定化酶的稳定性。结果表明:壳聚糖固定化木瓜蛋白酶的最佳条件为:壳聚糖浓度2.5%,加酶量0.3g/g载体,时间6h,温度15℃,pH7.5,酶活力回收率38.98%。提取牛蒡多糖的最佳条件为:固液比1:20,pH6.5,温度60℃,时间8h,加酶量1.8g/g载体,多糖提取率11.04%。固定化酶重复使用五次,酶活力仍保持50%以上。     

淘汰蛋鸡肉嫩化试验研究

采用木瓜蛋白酶,氯化钙对淘汰鸡肉进行嫩化处理,测定处理前后鸡肉的剪切力值、pH值及增水率变化,结果表明木瓜蛋白酶和CaCL2均有良好的嫩化效果。     

RADIOIMMUNOASSAY OF PAPAIN IN BEER

A radioimmunoassay (RIA) of papain is described which is capable of detecting 0·2 μg of papain/ml in beer, a level approximately 1% of that normally used for chillproofing. Changes in incubation conditions significantly accelerates the papain determination with only slight loss of accuracy. Apart from the high sensitivity it has been shown that the method is highly specific and distinguishes papain from the other chillproofing enzymes used.     

尿素对生丝木瓜蛋白酶脱胶的影响

将尿素与木瓜蛋白酶复配用于生丝脱胶,将生丝分别置于水、木瓜蛋白酶、尿素以及木瓜蛋白酶和尿素的混合溶液中在一定条件下进行脱胶处理,通过减量率的变化研究尿素对生丝木瓜蛋白酶脱胶的影响。结果表明：单独用尿素对生丝脱胶的减量低;尿素与木瓜蛋白酶复配用于脱胶,对木瓜蛋白酶没有毒害作用;在较高浓度下可有效提高生丝用木瓜蛋白酶脱胶的减量,使脱胶用酶量大幅降低,酶脱胶温度宽泛化。由于尿素在低浓度下对木瓜蛋白酶脱胶减量的提高基本没有作用,在高浓度下才能充分地体现其作用,要将其与木瓜蛋白酶复配用于脱胶,只有在大幅度降低其用量后才有可能,因此还需要进一步的研究。     

THE DETERMINATION OF PAPAIN IN BEER

The quantitative determination of papain in beer has been achieved by means of turbidity measurements on a casein substrate. The method, which is quick and simple, will measure activities of the order of 1–20 p.p.m. of crude papain. This will cover any amount likely to be found in beer     

Effects of Antemortem Injected Crude Papain in Chicken Muscle

The effects of antemortem injected crude papain in chicken muscle on muscle protein hydrolysis were investigated. White Leghorn hens were injected intravenously prior to slaughter with crude papain or heat denatured crude papain. Collagen solubility (hydroxyproline) at 60°C was 210% of the heat denatured crude papain control. SDS-PAGE revealed no difference between treatments. Crude papain was unable to penetrate intact cells at physiological temperatures. A small but significant increase in muscle TCA soluble tyrosine was observed due to antemortem papain treatment. Postmortem papain treatment resulted in a 14 fold increase in tyrosine. Little if any intracellular muscle protein hydrolysis occurred as a result of antemortem papain treatment. Collagen hydrolysis may be the major contributor to antemortem papain induced meat tenderness.     

木瓜蛋白酶水解核桃粕制备抗氧化活性多肽的研究

利用木瓜蛋白酶水解核桃粕,得到核桃多肽。采用Box-Behnken中心组合分析优化水解条件,以羟基自由基和超氧阴离子自由基清除率为响应值,得到最优水解条件为:pH值6、温度50℃、酶解时间3h、底物浓度3%。在此条件下,核桃多肽对羟基自由基和超氧阴离子自由基的清除率分别为32.7%和24.6%。结果表明,木瓜蛋白酶水解核桃粕得到的核桃多肽具有一定的抗氧化活性;并且与VC做对比,其还原能力是VC的37.9%。     

木瓜蛋白酶的提取及应用研究进展

本文主要介绍木瓜蛋白酶的组成,重点阐述提取方法、提取工艺的研究进展,进一步介绍木瓜蛋白酶在食品工业特别是在食品加工中的应用,并对其在医药、化工及未来食品加工方面的应用前景进行展望。     

Tetrathionate Protects Proteolytic Activity of Simulated Papaya Latex and Crude Papain

Sodium tetrathionate (TT) was assessed for its possible protection of proteolytic activity (PA) of rehydrated crude papain during drying and storage. The addition of 1% TT significantly decreased the loss of PA of rehydrated crude papain after being oven dried at 55°C. This protection of PA was 15–20% higher than that from metabisulfite. Crude papain with 1% TT lost 20% of PA and with metabisulfite lost 45% of PA in 13 wk at room temperature. Results suggest TT may decrease loss of PA during papaya latex processing and storage of crude papain.     

酶制剂对藜麦酚类物质及馒头品质的影响

为探究酶制剂对藜麦馒头中酚类物质、馒头色泽、弹性和口感等品质的影响及其对营养功能的作用效果,试验以藜麦为原料,采用木聚糖酶、真菌淀粉酶、果胶酶、木瓜蛋白酶酶解后混合30%谷朊粉制作藜麦馒头,测定质构、游离酚、结合酚、黄酮含量及抗氧化活性。结果表明：与对照组相比,酶处理的藜麦馒头物性数据均有所提升,其中真菌淀粉酶组的硬度、弹性、咀嚼性分别提高87%、8%、96%;木瓜蛋白酶组的硬度、弹性、黏聚性分别降低30%、20%、26%。在多酚含量方面的影响,木瓜蛋白酶组总酚含量提高99.23%,但果胶酶组结合多酚提高约32.67%。木聚糖酶组游离黄酮的含量增加25%,而结合黄酮含量降低17.98%。木瓜蛋白酶处理藜麦馒头的游离和总多酚提取物的DPPH自由基清除率分别增加96.67%、61.60%,而游离多酚铁离子还原能力增加32.30%。真菌淀粉酶增强藜麦馒头的硬度、弹性、咀嚼性,木瓜蛋白酶处理释放出更多的游离酚,木聚糖酶释放更多结合多酚,从而发挥抗氧化作用。     

超高压结合酶法消减南美白对虾蛋白过敏原研究

以南美白对虾为研究对象,提取水溶性虾蛋白,采用超高压法、超高压处理后再用木瓜蛋白酶水解、超高压下直接酶解等方法消减其过敏原蛋白,用间接酶联免疫吸附法检测过敏原消减效果,确定消减过敏原的条件。结果表明：采用超高压法处理,其最佳条件为压力150MPa、温度45℃、保压时间35min,产物抗原抗体反应的OD492nm值为0.1997;高压结合木瓜蛋白酶水解法最佳条件为：350MPa、温度45℃、保压时间20min,产物抗原抗体反应的OD492nm值为0.0492;超高压下直接用木瓜蛋白酶处理,其最佳条件为：压力300MPa、温度45℃、保压时间35min,产物抗原抗体反应的OD492nm值为0.05。由此可见,高压对过敏原蛋白有消减作用,先高压再水解和超高压下直接酶处理对过敏原的消减效果更好。     

Purification and determination of inhibitory activity of recombinant soyacystatin for surimi application

A recombinant soyacystatin (r-soyacystatin) was tested for its inhibitory activity against cysteine proteinase of Pacific whiting and its activity was compared to that of egg white cystatin. A recombinant soyacystatin expressed in Escherichia coli was purified to electrophoretic homogeneity using phenyl-Sepharose and DEAE-Sepharose. Native egg white cystatin was purified by using affinity chromatography on CM-papain-Sepharose generated in our lab. Egg white cystatin and soyacystatin were tested for proteinase inhibitory activity against commercial papain and also cathepsin L purified from Pacific whiting muscle. The r-soyacystatin exhibited papain-like protease inhibition activity comparable to that of the egg white cystatin, which could inhibit papain and Pacific whiting cathepsin L. The r-soyacystatin subsequently inhibited the autolytic activity of Pacific whiting surimi.     

超高压对木瓜蛋白酶构象及酶活力的影响

为研究超高压对木瓜蛋白酶构象变化与酶活力的影响,本研究利用红外光谱法和荧光光谱法对木瓜蛋白酶经超高压处理后的结构变化及特定氨基酸微环境变化进行了分析。结果表明：与常压相比,超高压处理对木瓜蛋白酶活力均有显著影响（P＜0.05）。其中,200 MPa、37 ℃、20 min处理时,酶活力在所选处理范围内达到最大,较常压下的酶活力提高了6.8%;红外光谱结果表明：超高压处理后,木瓜蛋白酶二级结构变化与酶活力变化相关性较差;荧光光谱结果显示：200 MPa、37 ℃处理木瓜蛋白酶20 min,228 nm波长激发后木瓜蛋白酶的外源性荧光强度达到最低（2 023）,荧光强度变化与酶活力变化规律有良好的相关性。因此,木瓜蛋白酶活性变化与疏水性氨基酸的暴露程度有关,暴露程度越小,酶活力越大。     

柔性固定化木瓜蛋白酶的制备及在酶解波纹巴非蛤中的应用

在壳聚糖膜的表面接上分子链较长的双醛淀粉作为柔性固定化酶的载体,将木瓜蛋白酶固定在该载体上制成柔性固定化木瓜蛋白酶,并应用于酶解波纹巴非蛤制备小分子肽。通过考察温度、pH 值、加酶量和固定化时间等条件对柔性固定化木瓜蛋白酶的酶活、酶活回收率的影响,确定最优固定化条件。结果表明：双醛淀粉用量0.8mg/g 壳聚糖、柔性固定化酶温度25℃、柔性固定化酶时间20h、加酶量40mg/g 壳聚糖、pH8.0 条件下所得的固定化酶酶活最高,酶活回收率63.35%。将该柔性固定化酶在酶解温度40℃、pH7.0、加酶量1.0%、酶解时间4h条件下制备波纹巴非蛤小分子肽,产率为3.4055%。     

Proteolytic activity and immunoreactivity of chemically modified papain

The effect of different chemical modifications of papain on the proteolytic activity and immunoreactivity has been studied. Modification with Dextran T 2000 caused increasing decline in proteolytic activity both with increasing degree of dextran oxidation and amount of bound dextran, whilst the immunoreactivity determined by nephelometry remained unchanged. Modification of papain with glutaraldehyde and formaldehyde causes rapid drops in both activities, even at very low concentrations of agents. Acetylation of papain showed expressive maxima of both proteolytic and immunochemical activities at the same degree of enzyme acetylation. Modification with diazobenzenesulfonic acid caused a high increase in immunoreactivity and a small increase in proteolytic activity.     

固定化木瓜蛋白酶的研究进展

木瓜蛋白酶具有水解蛋白质能力强的特点,但游离的木瓜蛋白酶稳定性差,难以回收利用;固定化木瓜蛋白酶既保持了酶的催化性质,又克服了游离酶的不足,具有广泛的发展前景。