Influence of lactic acid bacteria, cellulase, cellulase-producing Bacillus pumilus and their combinations on alfalfa silage quality

This study assessed the effects of lactic acid bacteria(LAB), cellulase, cellulase-producing Bacillus pumilus and their combinations on the fermentation characteristics, chemical composition, bacterial community and in vitro digestibility of alfalfa silage. A completely randomized design involving a 8(silage additives)×3 or 2(silage days) factorial arrangement of treatments was adopted in the present study. The 8 silage additive treatments were: untreated alfalfa(control); two commercial additives(GFJ and Chikuso-1); an originally selected LAB(Lactobacillus plantarum a214) isolated from alfalfa silage; a cellulase-producing Bacillus(CB) isolated from fresh alfalfa; cellulase(C); and the combined additives(a214+C and a214+CB). Silage fermentation characteristics, chemical composition and microorganism populations were analysed after 30, 60 and 65 days(60 days followed by exposure to air for five additional days). In vitro digestibility was analysed for 30 and 60 days. Compared with the other treatments, selected LAB a214, a214 combined with either C or CB, and Chikuso-1 had the decreased(P<0.001) pH and increased(P<0.001) lactic acid concentrations during the ensiling process, and there were no differences(P>0.05) among them. Fiber degradation was not significant(P≥0.054) in any C or CB treatments. The a214 treatment showed the highest(P=0.009) in vitro digestibility of dry matter(595.0 g kg^(–1) DM) after ensiling and the highest abundance of Lactobacillus(69.42 and 79.81%, respectively) on days 60 and 65, compared to all of other treatments. Overall, the silage quality of alfalfa was improved with the addition of a214, which indicates its potential as an alfalfa silage inoculant.

Effect of Cellulase and Lactic Acid Bacteria on Fermentation Quality and Chemical Composition of Wheat Straw Silage

The object of this study was to determine the effect of cellulase and lactic acid bacteria (LAB) on fermentation quality and chemical composition of wheat straw silage. Silages were prepared using a small-scale fermentation system and the moisture level was adjusted to 60% of fresh matter (FM) with deionized water. Treatments were designed as: control silage without additives, LAB inoculant Lactobacillus casei Z3-1 (1.0 × 106 cfu·g-1 of FM), commercial inoculant L. plantarum FG 1 (1.0 × 106 cfu·g-1 of FM), Z3-1 + cellulase and FG 1 + cellulase. The neutral detergent fiber (NDF), acid detergent fiber (ADF) and crude protein (CP) contents of the wheat straw prior to ensiling were 76.93%, 48.52% and 4.63% of dry matter (DM), respectively. After 30 days of fermentation, the silages treated with LAB and LAB + cellulase had a lower (P < 0.05) pH and higher (P < 0.05) lactic acid content than the control, and the coliform bacteria, yeast and mold were inhibited at the early stage of fermentation. Besides, silages treated with cellulase had lower (P < 0.05) values of ADF and NDF than the control. The results confirmed that the addition of cellulase and LAB contributed to improving the fermentation quality of wheat straw silage.

Production of Citric Acid from Apple Pomace Enzymolyzed by Cellulase

Cellulase can evidently increase the content of glucose and has a significant effect on the production of citric acid from apple pomace by Aspergillus niger. Based on experiments, a cellulolytic enzyme named cellulase A6 was found able to produce about 170 g glucose from 1 kg dried apple pomace after 12 h reaction, with cellulase concentration of 20 U/g in the medium at 50oC, natural pH without pretreatment of alkali. Using the treated apple pomace as a liquid state substrate, Aspergillus niger-C selected out was able to produce about 256 g citric acid from 1 kg dried apple pomace at 35oC in 3 d or 30oC in 5 d with flask rotation speed of 210 r/min, and the conversion of citric acid could reach 80% based on the amount of sugar consumed.

Study on Addition Amount of Lactic Acid Bacteria and Cellulase in Mixed Silage of Potato Plants

[Objectives]This study was conducted to investigate the appropriate addition amounts of lactic acid bacteria and cellulase in the mixed silage of potato plants,so as to provide a basis for the rational use of potato plants.[Methods]Fresh potato(Solanum tuberosum)seedling plants,watermelon(Citrullus lanatus)plants,and melon(Cucumis melo)plants were selected as raw materials for silage,each of which was added at a ratio of 32%,and 4%of brown flour was added additionally.On this basis,an L 9(32)orthogonal experiment was designed to investigate the effects of the amount of lactic acid bacteria(10,20,30 g/kg)and the amount of cellulase(5,10,15 g/kg)on the experimental results with sensory evaluation score as an investigation index.[Results]With the increase of the amount of lactic acid bacteria and the amount of cellulase,the sensory evaluation score of silage increased,and it was the highest when the amount of lactic acid bacteria added was 20 g/kg and the amount of cellulose was 10 g/kg.With the increase of the amounts of lactic acid bacteria and cellulase added,ammonia nitrogen showed a decreasing trend,and the pH and ammonia nitrogen were the lowest when the amount of lactic acid bacteria added was 20 g/kg and the amount of cellulose was 10 g/kg.[Conclusions]This study provides a theoretical basis for the rational use of effective ingredients and nutrients in potato plants and the development of new feed resources.

Cellulase Producing Bacteria from the Wood-Yards on Kallai River Bank

This study evaluates the influence of growth parameters such as pH, temperature, Carboxy Methyl Cellulose (CMC) concentration and agitation on cellulase production from three bacterial strains, viz., Achromobacter xylosoxidans BSS4, Bacillus sp. BSS3 and Pseudomonas sp. BSS2 isolated from the wood-yards on Kallai river bank in Kerala. Production of cellulase by these isolates was detected using basal salt medium (BSM) with 0.5% CMC as supplement, and CMCase activity was confirmed by iodine test. Dinitrosalicylic acid method was employed for assaying the cellulase production by measuring the amount of glucose liberated in μmol/mL/min. Maximum enzyme production from Pseudomonas sp. BSS2 was at pH 8, 37℃ with 1% CMC and 150 rpm, and cellulase production increased from initial 49.84 U/mL to 91.28 U/mL after optimization. The highest enzyme activity from Bacillus sp. BSS3 was at pH 9, 37℃ with 1% CMC, 150 rpm, and cellulase production increased from initial 26.05 U/mL to 104.68 U/mL after optimization. The maximum enzyme production from A. xylosoxidans BSS4 was at pH 7, 40℃ with 0.5% CMC and 150 rpm, and cellulase production increased from initial 55.28 U/mL to 68.37 U/mL after optimization. Thus among the three isolates, Bacillus sp. BSS3 showed maximum enzyme yield which can be explored for further scale up studies with an industrial perspective.

赖氨酸修饰对梯棱羊肚菌黑色素结构和理化特性的影响

为了进一步提高梯棱羊肚菌黑色素的提取率及溶解性,该研究采用单因素、Plackett-Burman试验、响应面试验对纤维素酶-超声波协同提取梯棱羊肚菌黑色素的提取工艺进行优化。通过赖氨酸修饰,并对修饰前后的梯棱羊肚菌黑色素进行结构表征、理化性质及稳定性分析。结果表明,在NaOH浓度为1.54 mol/L,纤维素酶添加量为20 mg/g,纤维素酶酶解时间为78.6 min,料液比为1:30,酶解温度为40℃,超声时间为80 min条件下提取的梯棱羊肚菌黑色素最优。未修饰的梯棱羊肚菌黑色素不溶于水,色价值为480.24,修饰后的黑色素溶解度为1 016 g/L,色价值为1 771.18,比未修饰的黑色素在溶解性、色价值方面均有所提高。此外,在不同的温度、光照、pH条件下,修饰前后的梯棱羊肚菌黑色素均比较稳定。以上研究结果为梯棱羊肚菌黑色素的高效提取及其产品的开发利用奠定了理论基础。

固态发酵改善棉籽粕品质的研究

通过固态发酵对棉籽粕进行品质改善,并探究其发酵后感官评价、抗营养因子含量及粗蛋白、总酸、酸溶蛋白的变化,为棉籽粕在水产养殖行业中的应用提供理论基础。研究首先探究了枯草芽孢杆菌(Bacillus subtilis)单菌发酵棉籽粕降解其抗营养因子含量的最佳接种量、水料比和时间。研究结果表明,当接种量达到11%为最佳,水料比在(0.6—0.8﹕1)最佳,当发酵时间达到60h后,各抗营养因子的降解率达到最高。随后采用酿酒酵母(Saccharomyces cerevisiae)、植物乳杆菌(Lactobacillus plantarum)配合枯草芽孢杆菌,同时在底物中添加植酸酶及纤维素酶溶液进行混菌固态发酵,并对最佳混菌比例以及酶液添加比例进行探究,结果表明,当枯草芽孢杆菌、酿酒酵母和植物乳杆菌的混菌比例为1﹕1﹕2时综合效果最佳,纤维素酶添加比例为1.25%,植酸酶添加比例为0.10%,降解抗营养因子的效果最佳。随后,以此条件进行混菌发酵,结果表明,混菌发酵棉籽粕在感官评定上要优于单菌发酵,棉籽粕中游离棉酚降解率达到40.13%,植酸降解率达到34.92%,单宁降解率达到40.24%,粗蛋白提升了10.26%,酸溶蛋白提升了59.2%,总酸提升了79.49%。因此,研究为提高棉籽粕的饲用价值提供参考,有利于提高棉籽粕在水产饲料中的添加水平。

Effects of lipoic acid on the activities of fibrinase, antioxidant enzymes and related factors in patients with type 2 diabetic foot

Objective:To investigate the effects of lipoic acid on the activities of fibrinase, antioxidant enzymes and related factors in patients with type 2 diabetic foot.Methods: 120 patients with type 2 diabetic foot treated in the Department of Endocrinology and Metabolism in our hospital from March 2015 to March 2018 were collected and randomly divided into control group and observation group, 60 cases in each group. The patients in control group were treated with alprostadil, and the patients in observation group were treated with lipoic acid combined with alprostadil. Before and after treatment, the levels of plasminogen activator, fibrinopeptide A, plasminogen activator inhibitor-1, superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), catalase (CAT), Toll-like receptor 2 (TLR2), Toll-like receptor 9 (TLR9), vascular cell adhesion molecule-1 (VCAM-1), matrix metalloproteinase-9 (MMP-9) and homocysteine (Hcy) of patients in the two groups were detected and compared.Results: After treatment, the levels of plasminogen activator, SOD, GSH-PX and CAT of patients in the control group and the observation group were significantly increased. The levels of fibrinogen A and plasminogen activator inhibitor -1 of patients in the control group and the observation group were significantly decreased. The changes of the above indexes of patients in the observation group were more obvious than those in the control group (P<0.05). The levels of TLR2, TLR9, VCAM-1, MMP-9 and Hcy in the serum of patients in the control group and the observation group were significantly decreased. The down-regulation of these factors of patients in the observation group was more obvious than that in the control group (P<0.05).Conclusion:Lipoic acid combined with alprostadil in the treatment of type 2 diabetes mellitus disease can significantly enhance the activity of fibrinolytic enzymes and antioxidant capacity, alleviate inflammation, and it has a good clinical effect.

蔗渣预处理发酵产纤维素酶研究

纤维素预处理可以使纤维素与木质素、半纤维素分开,从而提高酶类对纤维素和半纤维素的可及度,提高纤维素水解率。本实验采用蔗渣为原料,将其粉碎后分别用40、60、80、100目的筛子过筛得到各种粗细程度不同的蔗渣。然后用2%稀硫酸和10%氨水联合对其进行预处理,再以预处理过的蔗渣为碳源培养绿色木霉研究其产酶情况。先以10% NH_(4)OH处理,再用2% H_(2)SO_(4)的处理经发酵后滤纸酶活为6.90 U/mL,达到最高。先以2% H_(2)SO_(4)处理再用10% NH_(4)OH的预处理方法滤纸酶活为6.79 U/mL。经过预处理的蔗渣培养所产滤纸酶活力从高到低排列:(碱/酸)预处理>(酸/碱)预处理>酸预处理>碱预处理>未处理。在同种处理方法下过80目筛子所得的蔗渣所产的滤纸酶活力最高,其酶活力从高到低排列:80目>60目>40目>100目。

黑曲霉纤维素酶对蓝靛果酵素中挥发性化合物和有机酸的影响

为开发不同风味的蓝靛果酵素,对黑曲霉纤维素酶在蓝靛果酵素中的作用进行研究,在蓝靛果中接种0.5%的乳酸片球菌,添加或不添加黑曲霉纤维素酶进行发酵并用气质联用仪和高效液相色谱仪对产品进行分析。结果表明,添加黑曲霉纤维素酶后蓝靛果酵素(fermented blue honeysuckle berry cellulase,FBHB-C)的挥发性化合物共有44种,不添加黑曲霉纤维素酶的蓝靛果酵素(fermented blue honeysuckle berry,FBHB)有42种,两个样品的醇类化合物含量分别占各总挥发性化合物含量的75.95%和24.77%、酯类化合物占总挥发性化合物含量的4.38%和53.00%、酮类化合物占3.17%和11.38%、醛类化合物占14.06%和6.33%、其他化合物占2.44%和4.51%,FBHB-C主要挥发性化合物为顺式-4-己烯-1-醇、正己醇、1-辛醇、苯甲醛、仲辛酮、苯甲酸乙酯。在两种蓝靛果酵素中检出有机酸7种,柠檬酸含量最多,FBHB-C中草酸、酒石酸、苹果酸、富马酸、马来酸的含量比FBHB中相应有机酸分别提高130.38%、88.03%、196.55%、1509.52%、156.27%,其中草酸、酒石酸、柠檬酸呈香贡献最大。

纤维素酶、乳酸菌对干糜草发酵品质与营养价值的影响

试验旨在研究纤维素酶、乳酸菌等菌剂发酵糜草秸秆发酵品质和营养价值。试验设置7个处理组,分别为青干贮专用复合酶组(FG)、瑞森复合乳酸菌组(FR)、纤维素酶组(XM)、纤维素酶+复合乳酸菌组(XMR)、瑞森青贮宝组(MQ)、青干贮专用复合酶+复合乳酸菌组(FGQ)、对照组(MCK),每组3个重复。试验期60 d。结果显示,糜草含有一定的总黄酮和多酚氧化酶。XM组、MQ组感官评价为优等。XMR组蛋白质(CP)含量显著高于FG组和FGQ组(P<0.05);XMR组粗纤维(CF)、中性洗涤纤维(NDF)、酸性洗涤纤维(ADF)含量均极显著低于对照组、糜草原样(P<0.01)。MQ组糜草秸秆发酵后有机酸总和、乳酸占比分别极显著高于对照组(P<0.01);MQ组、XM组乙酸占比较合适;FGQ组、MQ组、XMR组、XM组丁酸含量极显著低于对照组(P<0.01);XM组、XMR组、MQ组、FGQ组综合得分在60~75分之间,评价为良好。研究表明,采用纤维素酶、青贮宝、纤维素酶+复合乳酸菌等处理糜草秸秆后,显著提高了糜草发酵品质,明显提高了营养价值和饲用价值;糜草中总黄酮含量明显升高,具有较高的开发利用价值。

Abscisic acid and ethephon regulation of cellulase in the endosperm cap and radicle during lettuce seed germination

The purpose of this study was to investigate the role of cellulase in endosperm cap weakening and radicle elongation during lettuce(Lactuca sativa L.) seed germination. The application of abscisic acid(ABA) or ethephon inhibits or promotes germination, respectively, by affecting endosperm cap weakening and radicle elongation. Cellulase activities, and related protein and transcript abundances of two lettuce cellulase genes, Ls CEL1 and Ls CEL2, increase in the endosperm cap and radicle prior to radicle protrusion following imbibition in water. ABA or ethephon reduce or elevate, respectively,cellulase activity, and related protein and transcript abundances in the endosperm cap. Taken together, these observations suggest that cellulase plays a role in endosperm cap weakening and radicle elongation during lettuce seed germination, andthat the regulation of cellulase in the endosperm cap by ABA and ethephon play a role in endosperm cap weakening.However, the influence of ABA and ethephon on radicle elongation may not be through their effects on cellulase.

西咪替丁片中微晶纤维素的测定研究

目的:建立一种测定制剂中微晶纤维素(MCC)的分析方法。方法:以纤维素酶将纤维素水解成还原糖,测定还原糖得到MCC的含量。结果:本实验所用纤维素酶的最适温度为50℃,最适pH值为4.8;方法学验证结果显示,MCC质量浓度在0.0445~0.0691 mg/mL间呈线性,R^(2)=0.9956;平均回收率为102.8%。结论:本法精确度高、专属性强,解决了在复杂体系中难以测定MCC的问题,在药物处方剖析方面有较大意义。

不同添加剂对干谷草秸秆发酵品质与营养价值的影响

试验旨在研究添加青干贮复合酶、纤维素酶、复合乳酸菌等添加剂对干谷草秸秆发酵品质及营养价值的影响。试验设7组,GQ组添加3 g/kg青干贮专用复合酶,GQR组添加3 g/kg青干贮专用复合酶+1 g/kg复合乳酸菌,GX组添加3 g/kg纤维素酶,GXR组添加3 g/kg纤维素酶+1 g/kg复合乳酸菌,GF组添加3 g/kg复合乳酸菌,GY组添加3 g/kg瑞森青贮宝,对照组(GCK组)不添加添加剂,每组3个重复。室温发酵60 d。结果显示,GX组、GXR组、GF组、GY组发酵谷草秸秆的粗蛋白含量极显著高于GCK组(P<0.01);GQR组发酵谷草秸秆粗灰分含量显著高于GCK组(P<0.05),GXR组、GX组发酵谷草秸秆粗纤维含量极显著低于除GY组外的其他组(P<0.01)。GY组乳酸含量、乳酸占总有机酸比例均极显著高于GCK组(P<0.01),氨态氮/总氮比值极显著低于GCK组(P<0.01);GY组、GQR组、GF组发酵谷草秸秆综合评分均在85分以上,质量为优。研究表明,添加青贮宝、纤维素酶、复合乳酸菌、纤维素酶+复合乳酸菌可明显提高谷草秸秆的营养价值,有效改善发酵品质,同时谷草中含有较高的多酚氧化酶和总黄酮,具有较高的开发利用价值。

草甘膦对桉树林地土壤酶活性的影响

为探明施用草甘膦对桉树林地土壤酶活性的影响,在广西壮族自治区国有维都林场三年生桉树林地随机选择8块样地开展了试验。2020年和2021年连续2年对其中4块样地喷施41%草甘膦异丙胺盐水剂清除杂草和矮灌木丛,另外4块样地采用人工除草作对照,其他管护方式相同。2022年4月采集土壤样品,测定其土壤酶活性。结果表明:与对照样地相比,施用草甘膦的样地各土层土壤过氧化氢酶活性均显著降低,0~5 cm、>5~10 cm、>10~20 cm和>20~50 cm土层过氧化氢酶活性分别降低了25.98%、20.82%、21.85%和20.14%;各土层酸性磷酸酶活性均较对照样地下降,0~5 cm、>5~10 cm、>10~20 cm和>20~50 cm土层分别降低了19.16%、22.47%、2.29%和3.79%,仅0~5 cm和>5~10 cm土层降低显著;各土层纤维素酶活性也均较对照样地降低,0~5 cm、>5~10 cm、>10~20 cm和>20~50 cm土层分别降低了27.01%、5.10%、4.17%和5.32%,仅0~5 cm土层降低显著;各土层土壤脲酶和蔗糖酶活性均较对照样地有一定程度的提高,但差异不明显。由此表明,在桉树林地施用除草剂草甘膦,会明显降低表层土壤过氧化氢酶、纤维素酶和酸性磷酸酶的活性,对林地的养分循环造成不利影响,降低土壤肥力。

DNS法测定纤维素酶活力最适条件研究

鉴于纤维素酶研究和生产中，纤维素酶活力测定和酶活力单位的表示方法很不统一的现象，本文研究和分析了３，５－二硝基水杨酸法测定羧甲基纤维素酶糖化力的实验条件，如：温度、ｐＨ、酶促反应时间、ＤＮＳ显色时间、波长等．实验表明：在５０℃，ｐＨ４．６条件下酶促反应时间５ｍｉｎ，ＤＮＳ显色５ｍｉｎ，于４９０ｎｍ处测定ＯＤ值比较适宜．

乳酸菌与纤维素酶对草甸草原天然牧草青贮品质的影响

旨在探讨不同添加剂对天然牧草青贮的影响,提高天然牧草青贮的利用效率,指导天然牧草的生产与利用。本研究以草甸草原盛花期的天然牧草为材料,设置6个处理组,分别为SLAB(真空袋,乳酸菌(2×10~5cfu·g^(-1)))、SCE(真空袋,纤维素酶(0.8g·kg^(-1)))、SCK(真空袋,无添加,对照)、BLAB(青贮窖,乳酸菌(2×10~5cfu·g^(-1)))、BCE(青贮窖,纤维素酶(0.8g·kg^(-1)))、BCK(青贮窖,无添加,对照),每组3个重复,青贮60d后对不同添加天然牧草青贮饲料的微生物组成、发酵品质、营养成分进行比较分析。结果表明,所有添加剂组青贮效果均较好。添加乳酸菌和纤维素酶均可显著提高天然牧草青贮的乳酸菌含量、乳酸含量和乳酸/乙酸含量(P<0.05),青贮乳酸菌组乳酸菌含量高于纤维素酶组,添加剂提高蛋白质保存率(P<0.05),显著提高可溶性碳水化合物利用率(P<0.05),显著降低pH、氨态氮/总氮和酸性洗涤纤维含量(P<0.05),显著降低酵母菌含量(P<0.05),对灰分含量无显著影响(P>0.05)。综上表明,乳酸菌更适于用作天然牧草青贮饲料的添加剂,可在草原牧区将天然牧草由小包装青贮向大规模青贮推广应用。

添加剂对羊草青贮发酵品质和体外消化率的影响

研究乳酸菌制剂、纤维素酶和甲酸对羊草(Leymus chinensis)青贮发酵品质和体外消化率的影响。结果表明:添加乳酸菌制剂、纤维素酶、乳酸菌制剂+纤维素酶均显著降低了青贮饲料pH值,提高了乳酸含量(P<0.05);添加甲酸显著降低了青贮料pH值、乳酸和氨态氮含量(P<0.05);添加乳酸菌制剂或甲酸显著提高了干物质体外消化率(P<0.05),乳酸菌制剂与纤维素酶混合添加显著提高了干物质和粗蛋白质体外消化率(P<0.05)。

酶酸结合法降解植物纤维素新工艺

以麦秸为材料,研究乙酸、盐酸等对纤维素酶降解纤维素的影响;设计出降解纤维素的2个工艺方案:方案1是先酸解后酶解;方案2是先酶解后酸解。在这2种工艺方案基础上,研究一种酶酸循环降解法(CDCA法)。研究结果表明:方案2优于方案1;CDCA法亦明显优于方案1和方案2;以CH3COOH,HCl和纤维素酶为降解剂,在常压、温度低于100℃、反应12～15 h的条件下,CDCA法能使纤维素转化成葡萄糖的转化率达98.68%,反应终液中葡萄糖质量分数达5.38%;与常规技术相比,CDCA法只需25%的醋酸、50%的盐酸和少量的酶,可以大幅度地降低纤维素转化成葡萄糖的成本;本研究的酶酸结合法降解纤维素新工艺具有条件温和、工艺简单、成本低和周期短等特点,该工艺方法可为发酵工业上以纤维素为原料从事乙醇、抗生素、SCP等产品的开发生产及环保上固态生活垃圾的生物处理提供科学依据。