木质素高效降解菌的筛选及研究

(目的)筛选出可高效降解棉杆木质素的菌株;(方法)通过平板鉴定法进行初筛及复筛,再由固态发酵培养方式测定菌株对棉杆降解能力,并对棉杆木质纤维素含量的降解率及木质素酶活进行测定;(结果)筛选得到两株可高效降解棉杆木质素的菌株M11,H-1。H-1对棉杆木质素降解率的最大值为47%。而M11对棉杆木质素降解率的最大值为11%;(结论)棉杆木质素降解菌的研究为新疆棉杆的充分利用打下坚实的基础。     

不同云芝菌株腐朽杨木过程的扫描电镜研究

本文应用扫描电镜观察研究了白腐菌云艺（Coriolusvericolor）三个菌株腐朽杨木的过程，对试材的显微形态变化以及各菌株的降解特性进行了详细描述和探讨．实验结果表明，三个菌株降解木材的性能和方式存在着差异。NFU008对木质素和纤维素均具有很强的降解能力，它在腐朽早期优先降解胞间层中的木质素并能使纤维解离，在腐朽后期，它对纤维素产生强烈降解。NFU006对木质素和纤维素的降解是同时进行的，但对纤维次生壁中木质素的降解率明显高于对纤维素的降解率。NFU019降解木质素的能力相对较弱，但对纤维素的降解能力却很强，早期它以降解出生壁的木质素为主，而在腐朽后期它对纤维素产生剧烈降解。三个菌株最终均造成纤维细胞壁不同程度的减薄和破坏。     

木材腐朽真菌定向降解木质素为苯系芳香类化学品

对2种木材腐朽真菌降解木质素为一系列苯系芳香类化学品进行了初步研究,并与黄孢原毛平革菌进行了比较。从木材腐朽土壤样品中分离获得具有降解木质素活性的真菌11株,将其中活性最强的菌株ZS1与ZSH用于降解工业碱木质素与中性木质素。通过发酵条件的优化、降解产物种类与浓度的HPLC分析,结果发现,在初始底物浓度1.0～20.0g/L变化时,ZS1对碱木质素的降解率达到40.7%,比ZSH的大;而ZSH对中性木质素的降解率为32.3%,与黄孢原毛平革菌的降解能力相当,但比ZS1的大些。木质素能被定向降解为香草酸、苯甲酸、香草醛、苯酚、紫丁香醛、苯、对甲基苯甲醚、对甲基苯乙醚、甲苯与对二甲苯等芳香类化学品,而且不同真菌降解不同木质素的产物种类数量与浓度有所差异。这说明不同真菌对木质素的降解具有一定的底物依赖性,底物结构差异影响了降解率不同与产物种类差异,而且不同真菌降解木质素具有一定的产物定向选择性。     

一株木质素降解细菌的筛选及其降解途径

经高温碱性盐法预处理的玉米秸秆预处理液中木质素和碱性盐含量高,为了反复利用含盐预处理液,需脱除其中的可溶性木质素。本实验从腐木中筛选分离到一株木质素降解菌株CCZU-WJ6,通过形态学观察和16S rDNA序列分析,鉴定该菌株属于无色杆菌属（Achromobacter sp.）。测定了CCZU-WJ6木质素降解相关酶的活性,及其在玉米秸秆预处理液中的生长趋势、预处理液的COD去除率和木质素降解率,通过GC-MS和FTIR检测预处理液中组分及其化学键的变化。通过单因素实验确定了菌株CCZU-WJ6处理预处理液的最佳条件为pH 7.0、温度30℃、预处理液稀释倍数10倍、接菌量0.5g/mL。在处理6天后,预处理液COD脱除率为40.9%,木质素的降解率为32.1%,CCZU-WJ6分泌木质素过氧化物酶和锰过氧化物酶的酶活分别达到215.5U/L、200.1U/L。CCZU-WJ6会破坏木质素结构中的苯环结构、醚键以及C＝O键,降解产物中含有愈创木酚和对香豆酸,推断其降解途径为β-芳基醚代谢途径和阿魏酸代谢途径。CCZU-WJ6可用来处理含木质素的工业废水,在工业方面具有广阔的应用前景。     

选择性降解木质素白腐菌筛选的研究

对国内外15种产漆酶的白腐菌进行初步筛选,从中获得7株产漆酶活性较高的菌株。经对木材降解能力的分析及在培养过程中所产各种酶酶活的测定,得到了2株产漆酶酶活高、降解木质素选择性优良的菌株,其中Flammulinavelutipes和My-1两菌株经14天液体培养,漆酶酶活高达214IU/mL和231IU/mL。杨木经其处理30天后木质素降解率达到22.67%和15.46%。     

原生质体复合诱变选育刺芹侧耳木质素降解酶高产菌株

通过对刺芹侧耳(Pleurotus eryngiiGIM5.280)原生质体开展复合诱变及传代培养,以期得到遗传稳定的木质素降解酶高产菌株。结果表明,通过25s紫外诱变刺芹侧耳原生质体,正突变率达16%。对紫外诱变正突变株进行60Coγ二次复合诱变再生,获得五株正突变突出菌株。经过摇瓶发酵实验,发现这五株菌株产木质素降解酶能力较出发菌株明显提高。同时开展高产菌株的传代培养,检验其传代稳定性。连续传代四代测试结果表明,007号和167号菌株遗传的稳定性表现突出,发酵液中木质素降解酶产量稳定。尤其是007号菌株产酶可达到110U·mL-1,比出发菌株的酶活表达量提高54.3%。复合诱变能明显提高刺芹侧耳产木质素降解酶能力。     

不同培养条件对云芝(Coriolus versicolor)木质素降解酶产酶影响的研究

本文研究了不同培养条件对云芝（Coriolusversicolor）三种木质素降解前产酶的影响，发现三种木质素降解酶即水质素过氧化物酶（Lip）、赖锰标木质素过氧化物酶，（Mnp）和漆酶的产生和培养条件关系很大，添加黎芦醇（VOH）能显著地提高三种酶的产量，当VOH浓度为1mmol／L时，可使木质素过氧化物酶产量提高10倍，添加Mn2＋对赖锰木质素过氧化物酶有促进作用，但对木质素过氧化物酶有抑制作用。提高其它微量元素（Cu2+，Fe2+,Zn2+，Ca2+）等同样能显著促进三种酶的产生。采用30信浓度时，赖锰木质素过氧化物酶的量要比1培浓度时提高6倍。三种酶的最适产酶pH相差很大，木质素过氧化物酶在pH4．5～5．5左右，赖锰木质素过氧化物酶在PH5．5～6.5左右，而漆酶则抵达pH3．5～4．0．Tween80在低浓度时（0.01％-0.02％）有一定的促进作用，而在高浓度则有抑制作用。C／N比则是影响产酶的另一因素，低碳培养基比低氮培养基更适宜三种木质素降解酶的产生。同时，在低氮培养基和低碳培养基下，上述因素对三种酶产生的影响也存在显著差异。这有可能是云芝的木质素降解酶在不同的C／N下其产生机理存在差异。     

低温等离子体降解木质素磺酸钠的研究

将木质素磺酸钠水溶液通过空气气动法雾化,采用介质阻挡放电低温等离子体对其中的木质素磺酸钠进行降解。以紫外可见光谱(UV)、红外光谱(FTIR)和1H NMR谱等对降解前后的木质素磺酸钠进行了表征,结果表明,利用本实验方法降解木质素磺酸钠可行。实验同时考察了电极间隙、输出电压、催化剂用量、反应时间等工艺条件对木质素磺酸钠降解率的影响。实验结果表明,以硫酸铜为催化剂,在电极间隙为2 cm、输出电压为3 000 V、频率为10 kHz、反应时间为2.3 ms、催化剂用量为3.2%的条件下,质量浓度为1 g/L的木质素磺酸钠的降解率可达50%。     

酶解秸秆残渣中木质素的提取、表征和应用

采用碱性乙醇法从酶解玉米秸秆残渣原料中提取木质素，考察了乙醇浓度、温度、固液比和碱浓度对木质素提取率的影响；对提取的木质素进行了FTIR、13C NMR及1H NMR等表征，并探索了木质素部分替代聚己内酯二醇用于聚氨酯薄膜的合成及降解性能研究。研究发现，在50%乙醇浓度、200℃、m(固)∶m(液)=1∶20和0.5% NaOH浓度条件下，提取产物中木质素含量为94.1%，木质素提取率达到93.5%；产物的羟基含量为66.5 mg KOH/g，当用其添加27%质量分数的聚己内酯二醇制备聚氨酯薄膜时，薄膜的拉伸强度为24.0 MPa，断裂伸长率为224%，降解率为3.3%；进一步，添加适量葡萄糖提高聚氨酯薄膜的降解性能和耐水性能，降低薄膜透气性，葡萄糖添加量为3%时薄膜60天内的降解率能达到11%。     

高沸醇木质素及其衍生物对菠萝蛋白酶的吸附

通过曼尼希反应和室温相分离方法合成了高沸醇木质素胺和高沸醇木质素酚衍生物,并探讨了它们对菠萝蛋白酶的吸附特性. 结果表明,引入了胺基和酚羟基的高沸醇木质素衍生物对菠萝蛋白酶的吸附性能有很大的提高,吸附能力为HBS木质素胺>HBS木质素酚>HBS木质素,而且吸附后的菠萝蛋白酶活性仍保持在较高的水平,高沸醇木质素衍生物有望成为菠萝蛋白酶的浓缩吸附剂或固定化的载体.     

Comparative Biodegradation of Detergents I: Comparative Bacterial Biodegradation of Lauryl Sulphate and Tridecyl Benzenesulphonates

Twenty strains of bacteria isolated in a medium with lauryl sulphate as carbon source and twenty strains isolated with tridecyl benzenesulphonates were compared, with respect to growth and ability to degrade detergents, in submerged cultures containing one of these surfactants as major source of carbon. No strain was ideally suited for detergent biodegradation. Most bacteria isolated on lauryl sulphate rapidly degraded lauryl sulphate completely whereas they degraded tridecyl benzenesulphonates to a lesser extent than bacteria isolated on tridecyl benzenesulphonates. The latter strains, in turn, degraded lauryl sulphate at a lower rate and incompletely.     

Location of Lignin-Bonded Pectic Polysaccharides

The hypothesis that galactose and arabinose pectic substances are disributed with lignin throughout wood cell walls was tested by analyzing a high-yield loblolly pine kraft pulp that had been separated into middle-lamella-rich and secondary-wall-rich fractions. The fractions were first treated with a polysaccharidase enzyme mixture to obtain “enzyme lignins.” The structures of the galactose and arabinose oligomers remaining attached to the lignin were determined by methylation analysis. The 1→4 linked galactose structure, characteristic of pectic materials, was present in significant amounts in the middle-lamella-rich fraction, but to only a very small extent in the secondary-wall-rich fraction. In contrast, the pectic arabinan 1→5 linked structure was a significant proportion of the arabinose from both fractions. The 1→5 linked arabinose structure can arise from a linkage of arabinoxylan to lignin through the 5 position of the arabinose substituent as well as from pectic structures. After partial acid hydrolysis, approximately half the remaining arabinose in the secondary-wall-rich fraction was attached as monomeric units to lignin through the 5 position.     

Investigation of formaldehyde-free wood adhesives from kraft lignin and a polyaminoamide–epichlorohydrin resin

A formaldehyde-free wood adhesive system consisting of kraft lignin and a polyaminoamide-epichlorohydrin (PAE) resin (a paper wet strength agent) has been investigated in detail. The lignin-PAE adhesives were prepared by mixing an alkaline kraft lignin solution and a PAE solution. Mixing times longer than 20 min had little impact on the shear strength of the wood composites bonded with the lignin-PAE adhesives. The shear strength of the wood composites bonded with the lignin-PAE adhesives increased and then flattened out when the press time and the press temperature increased. The shear strength and water resistance of the wood composites bonded with the lignin-PAE adhesives depended strongly on the lignin/PAE weight ratio. Of the weight ratios studied, the 3:1 lignin/PAE weight ratio resulted in the highest shear strength and the highest water resistance of the resulting wood composites. The wood composites bonded with the lignin-PAE adhesives did not delaminate and retained very high strengths even after they underwent a boiling-water test. The lignin-PAE adhesives could be stored at room temperature for two days without losing their adhesion ability. PAE was the crosslinking agent in this lignin-PAE adhesive. Possible reactions between lignin and PAE are discussed in detail.     

Lattice Conversion of Cellulose in Wood

When peracetylated wood, prepared in an acetic anhydride-pyridine medium, was saponified and recrystallized, almost no conversion of cellulose I to cellulose II took place. Neither was conversion detected, when thirteen species of wood, differing in density from 0.17 to 0.71, as well as in lignin content from 18.1 to 38.8 %, were treated with 17.5 % aqueous NaOH, followed by recrystallization. The corresponding treatments for cellulose triacetate or cellulose resulted in complete conversion to cellulose II. Partial delignification of wood preceding the alkali-induced mercerization was found to cause partial lattice conversion to cellulose II, and the ratio of lattice conversion (L.C.R.) increased with the degree of delignification. The L.C.R. value reaches slightly more than 50 %, when one third of the lignin is removed, and the removal of about two third of the lignin from wood results in an almost complete conversion. The effect of lignin removal on lattice conversion was similar among the wood species, irrespective of their density and lignin content, Introduction of acyl groups larger than caproyl into wood, followed by saponification and mercerization resulted in a high conversion of cellulose in wood from cellulose I to cellulose II.     

固定化白腐菌对造纸废水的生物降解研究

就具有较强木质素降解能力的白腐菌对造纸废水的降解效果及条件进行了初步研究.分别采用固定化白腐菌和悬浮态白腐菌在不同接种量下对造纸废水进行降解,对降解过程中的白腐菌生长量、pH、COD、木质素含量等废水降解指标进行测定.结果显示两种不同状态下的白腐菌均能降解造纸废水,但它们的降解程度不同.其中固定化白腐菌降解木质素效果较好,且呈现较明显规律,说明固定化的白腐菌较悬浮态的白腐菌更具有降解造纸废水的潜在能力.     

油溶性表面活性剂生物降解度的测定

对ISO 10634-1995“水质———水介质中用于难溶有机物生物降解性能评价水溶液的制备与处理指南”推荐的方法进行逐一筛选,以期建立油溶性表面活性剂生物降解度测定的标准方法。实验结果表明,将油溶性表面活性剂超声乳化后,制备成粒径200 nm~300 nm乳液后,立即分散于水体系中进行降解,结果重复性较好,并且不影响油溶性表面活性剂生物降解度。用该方法对常见的油溶性表面活性剂的生物降解度进行了测定,结果表明大部分油溶性表面活性剂具有很好的生物降解性能;其降解规律与水溶性表面活性剂类似,疏水链长决定油溶性表面活性剂的生物降解性能,而亲水基团影响其生物降解速度。     

Formic acid/acetone‐organosolv pulping of white‐rotted Pinus radiata softwood

Organosolv pulping of fungally pretreated samples of Pinus radiata was evaluated. A screening study using five white‐rot fungi indicated that Ceriporiopsis subvermisopora and Punctularia artropurpurascens were the most selective ones for lignin degradation. These fungi were further cultured in bioreactors containing 2.5 kg of wood chips. Fungally‐pretreated samples were delignified by formic acid/acetone (7:3) at 150 °C. Pulping kinetics and strength properties of the resulting unbleached pulps were evaluated. Delignification rates and xylan solubilization rates were higher for the decayed samples than for the undecayed control, except for the sample biotreated with P artropurpurascens for 30 days. C subvermispora proved appropriate for treating the wood samples before organosolv pulping, since pretreatment with this fungus resulted in faster wood delignification and pulps with lower residual lignin. Increases in tensile index ranging from 3% to 22% were observed for most pulps prepared from biotreated samples, independently both of the fungal species used in the pretreatment and of the extent of the wood biodegradation expressed as wood weight loss. However, tear and burst indexes and brightness were lower than or similar to those of pulps prepared from the undecayed control. © 2000 Society of Chemical Industry     

How Well Do MWL and CEL Preparations Represent the Whole Hardwood Lignin?

Hardwood lignin preparations were isolated using classical milled wood lignin (MWL) and cellulolytic enzyme lignin (CEL) protocols. Furthermore, we managed to produce a lignin preparation of a very high yield, above 90%, with high purity and minimal structural degradation. This was achieved by dissolution and regeneration of milled wood prior to enzymatic hydrolysis, along with the use of 80% dioxane for lignin extraction. This preparation (RCEL-80) yield was about 4.5 and 2.5 times higher than the yields of the traditional MWL and CEL preparations, correspondingly, at the same milling time. The preparations obtained were comprehensively analyzed with state-of-the-art quantitative NMR techniques and wet chemistry methods. CELs were representative preparations for hardwood lignins as the S/G ratios in the CELs were equivalent to those in lignin in situ. Degradation of the main lignin subunits was very low. Importantly, the structures of CELs were independent of the preparation yield and were very similar to the structure of the high-yield lignin, RCEL-80. In contrast, the structures of MWLs were noticeably dependent on the preparation yield, especially when the yield was below 15–20% of total wood lignin. In particular, the S/G ratio increased with increasing MWL yield, but was still lower than the S/G ratios of the whole lignins in situ, even at high MWL yields. The amounts of β-O-4 lignin units in MWL were lower than those in the corresponding CEL preparations. It has been concluded that CEL preparations were representative of the whole wood lignin whereas MWLs represent a fraction with a lower S/G ratio.