Chemical composition and biodegradability of crop residues colonized by white-rot fungi

Two experiments were conducted to study the degradation of rice (Oriza sativa L) straw and maize (Zea maize L) stover by selected white-rot fungi, using loss of organic matter, disappearance of cell wall constituents, and in-vitro dry matter digestibility (IVDMD) as indices of rumen microbial degradation. Four white-rot fungi—Cyrathus stercoreus, Dichomitus squalens and two strains (cellulase-less mutant and a wild type) of Phanerochaete chrysosporium—were selected for the first experiment. After 30 days of solid state fermentation. C stercoreus increased IVDMD of both substrates from 373 g kg^?1 (rice) and 393 g kg^?1 (corn) to approximately 540 g kg^?1 by selectively degrading hemicellulose as opposed to cellulose. The highest IVDMD was obtained with a minimal loss of dry matter (33 g kg^?1). In contrast, both strains (wild and mutant) of P chrysosporium degraded cellulose and hemicellulose indiscriminately, and thus lowered the IVDMD to 276. 257 as opposed to 373 for rice straw and 301. 307 as opposed to 393 g kg^?1 for maize stover. All the fungi showed a preference for degrading maize stover compared with rice straw, although the materials were similar in cellulose and lignin concentration. No direct correlation was found between lignin degradation and the improvement of substrate IVDMD of either substrate. The objective of the second experiment was to determine the rate and extent of digestibility of rice straw colonized by C stercoreus during two incubation periods (15 and 30 days). Dry matter disappearance (DMD) was determined by ruminal incubation of the substrate contained in nylon bags. The 30-day incubation period was superior (1.3 ×) to that of the 15-day period in improving in-situ DMD. Treatment of rice straw for 30 days not only improved the extent of dry matter digestibility (616 as opposed to 389 g kg^?1), but also increased (1.5 ×) the rate of disappearance of substrate.     

Structural changes in wheat straw components during decay by lignin-degrading white-rot fungi in relation to improvement of digestibility for ruminants

Structural changes and resulting in-vitro dry matter digestibility (IVDMD) were determined during the course of solid-state fermentation of wheat straw using the lignin-degrading white-rot fungi Sporotrichum pulverulentum, Pycnoporus cinnabarinus and Cyathus stercoreus. The first fungus grew very rapidly on straw but degraded hemicelluloses and cellulose non-selectively resulting in very low IVDMD increases (50% after 35% weight loss). P. cinnabarinus and C. stercoreus preferentially degraded hemicelluloses achieving high improvements in IVDMD (maximum increase of 63 and 94%, respectively) with limited dry weight losses (12 and 18% after 7 and 13 days, respectively). The three fungi exhibited some selectivity among the individual hemicellulose components: O-acetyls were removed essentially at the same rate as xylan, while uronic acids accumulated as incubation proceeded. Conversely, the arabinose content decreased rapidly, especially with C. stercoreus and P. cinnabarinus, suggesting that removal of this pentose was partly responsible for digestibility improvement. Esterified phenolic acids were rapidly degraded during the first stages of decay by all three fungi although P. cinnabarinus and C. stercoreus degraded ferulic acid faster than p-coumaric acid. Lignin was preferentially degraded compared to polysaccharides by all three fungi. The amount of lignin removed, as determined by Klason, correlated well with IVDMD improvement (r=?0.97), while acid detergent lignin (ADL) showed a lower correlation (r=-0.86). Acidolysis yields of decayed lignin pointed to preferential degradation of β-O-4 ether linked units by the fungi. Syringyl units were removed faster than guaiacyl units only after 5 to 10% weight loss was obtained.     

Biodegradation of cell wall components of maize stover colonized by white-rot fungi and resulting impact on in-vitro digestibility

Treatment of crop residues with some species of white-rot fungi can enhance digestibility. This study was conducted to investigate changes in in-vitro dry matter disappearance (IVDMD) and degradation of cell wall constituents in maize (Zea maize L) stover treated with three white-rot fungi: Cyathus stercoreus, Phlebia brevispora and Phanerochaete chrysosporium. Solid fermentation of maize stover for 28 days at 27°C improved IVDMD from 409 g kg^?1 (control) to 514 g kg^?1 for P brevispora and 523 g kg^?1 for C stercoreus. In contrast, growth of P chrysosporium reduced IVDMD from 409 to 298 g kg^?1. All fungi degraded cell wall p-coumaric acid (PCA) and ferulic acid (FA), but P chrysosporium was the least effective in degrading PCA and FA. Conversely, P chrysosporium degraded lignin 1·6 times more effectively than C stercoreus and 1·4 times more than P brevispora, indicating that lignin degradation alone cannot account for the IVDMD enhancement and that degradation of PCA and FA may be important. Hemicelluloses were preferentially and highly utilized by all the fungi. Cellulose was extensively degraded only by P chrysosporium (69% lost after 28 days of incubation), while substrate colonized by the other two fungi retained more than 84% original cellulose. Incubation of C stercoreus and P brevispora decreased the concentrations of both xylose and arabinose, but increased glucose concentration, whereas P chrysosporium removed less xylose and decreased glucose concentration. Preferential removal of arabinose over xylose by the fungi caused an increase in the xylose to arabinose ratio of the treated residues. Enhanced digestibility may have resulted from cleavage of lignin-carbohydrate bonds. Results of this study suggest that digestibility enhancement of maize stover colonized by white-rot fungi is regulated by a complex combination of various factors, including the degradation of structural carbohydrates, cell wall phenolic acids and lignin.     

Effects of sodium hydroxide on cereal straws in relation to the enhanced degradation of structural polysaccharides by rumen microorganisms

The action of sodium hydroxide on the rate and extent of loss of wall polysaccharides from wheat and barley straws incubated in the rumen of the sheep was studied using a nylon bag technique. Cellulose loss reached a maximum in straws treated with 10 g NaOH 100 g^?1 straw. Further increase in amounts of alkali applied resulted in additional solubilisation of straw constituents, particularly xylan, but did not further enhance cellulose degradation. Rates of degradation of isolated straw, cotton and wood celluloses correlated with three of four measures of cellulose crystallinity obtained by X-ray diffraction and infrared analysis. Treatment of cellulose samples with 20 g NaOH 100 g^?1 cellulose did not affect rates of loss or produce detectable changes to cellulose morphology. Loss of acetic acid esters, which accompanied alkali treatment, did not relate to improvements to digestibility. The rate of loss of cellulose from delignified neutral detergent fibre (NDF) was that shown by the isolated straw cellulose, despite the presence of acetylated hemicellulose. Deacetylation of delignified NDF with NaOH or sodium methoxide did not enhance rates of xylose or cellulose loss. In lignified wall material (NDF) the molar ratio acetyl:xylose (approximately 1:3.5) and proportion of the major phenolic acids (p-coumaric and ferulic acids) remained essentially constant throughout the course of degradation in the rumen. It is suggested that loss of phenolic acids, while not directly contributing to digestibility enhancement, may model the susceptibility of ester linkages formed between polysaccharide and polyphenolic material to alkali. The extent of solubilisation of p-coumaric acid produced by NaOH showed a linear relationship with cellulose digestibility. An upper limit of 40% acid detergent lignin and 55% of total phenolics were released by NaOH, the extent of release closely paralleling cellulose digestibility. Phenolic material resistant to alkali remained associated with wall polysaccharides and was not released from nylon bags until over 50% of cellulose (and hemicellulose) had been degraded.     

The effect of physical and chemical treatments on the degradation of wheat and barley straws by rumen liquor-pepsin and pepsin-cellulase systems

Samples of wheat and barley straw, milled through a 1 mm sieve, were ball milled to <0.25 mm as a representative physical treatment, or delignified by the acidchlorite procedure as a representative chemical treatment. The original and treated samples were examined using in-vitro digestion systems based on rumen liquor-pepsin and pepsin-cellulase. The digestibility was determined as well as the extent of digestion of the cellulose, hemicelluloses and lignin. The rumen liquor-pepsin system was able to digest a greater proportion of the original and treated straws than the pepsin-cellulase system while ball milling improved the digestibility more than delignification, irrespective of the digestion system. Core lignin was not digested by either system. Ball milling improved the digestibility of the cellulose fraction to a greater extent than the hemicellulose fraction, whereas delignification had a greater effect on hemicellulose digestion than on cellulose digestion. There were also differences in the rate of removal of the different sugar residues present in the hemicellulose fraction.     

Disappearance of cell wall monomeric components from fractions chemically isolated from alfalfa leaves and stems following in-situ ruminal digestion

Alfalfa (Medicago sativa L) leaves and stems were incubated in the rumen of an alfalfa-fed steer for 0, 12, 24 and 48 h. Residues were analysed for monosaccharides remaining in chemically isolated fractions. Pectic substances were rapidly and extensively degraded. Xyloglucans also were quite degradable, whereas glucuronoxylans were very resistant to fermentation. Cellulose disappearance was intermediate between that of xylan and xyloglucan. For all non-pectic polysaccharides, leaf material was more degradable than stem, and this difference between leaf and stem was greater for cellulose than for most hemicellulosic fractions. Acetyl groups associated with the oxalate-insoluble fractions were degraded similarly to xylans. Alkali-labile phenolic acid content was lower in stems than in leaves, but contributed only 4 g kg^?1 of leaf DM. Less than 10% of alkali-labile phenolics were associated with hemicelluloses. Structures which limit alfalfa xylan fermentation may include acetyl groups and linkages of uronic acid side chains to phenolic material.     

Evaluation of pretreatment with Pleurotus ostreatus for enzymatic hydrolysis of rice straw

The effects of biological pretreatment of rice straw using four white-rot fungi (Phanerochaete chrysosporium, Trametes versicolor, Ceriporiopsis subvermispora, and Pleurotus ostreatus) were evaluated on the basis of quantitative and structural changes in the components of the pretreated rice straw as well as susceptibility to enzymatic hydrolysis. Of these white-rot fungi, P. ostreatus selectively degraded the lignin fraction of rice straw rather than the holocellulose component. When rice straw (water content of 60%) was pretreated with P. ostreatus for 60 d, the total weight loss and the degree of Klason lignin degraded were 25% and 41%, respectively. After the pretreatment, the residual amounts of cellulose and hemicellulose were 83% and 52% of those in untreated rice straw, respectively. By enzymatic hydrolysis with a commercial cellulase preparation for 48 h, 52% holocellulose and 44% cellulose in the pretreated rice straw were solubilized. The net sugar yields based on the amounts of holocellulose and cellulose of untreated rice straw were 33% for total soluble sugar from holocellulose and 32% for glucose from cellulose. The SEM observations showed that the increase in susceptibility of rice straw to enzymatic hydrolysis by pretreatment with P. ostreatus is caused by partial degradation of the lignin seal. When the content of Klason lignin was less than 15% of the total weight of the pretreated straw, enhanced degrees of enzymatic solubilization of holocellulose and cellulose fractions were observed as the content of Klason lignin decreased.     

Effect of delignification on the in vitro rumen digestion of polysaccharides of bagasse

Sugar cane bagasse was partially delignified by three different chemical methods, viz. sodium sulphide/hydroxide (Kraft process), sodium hydroxide and sodium sulphite. The Kraft and alkali treatments each reduced the lignin content by about 55%, and increased the rumen dry matter digestibility (d.m.d., measured by an in vitro method) by over 170%. Treatment of the bagasse with sodium sulphite removed only 15% of the lignin and increased the d.m.d. by only 37%. The digestibility of hemicelluloses (measured as xylan and arabinan) and cellulose in delignified bagasse increased fourfold and twofold, respectively, for the Kraft and alkali treatments and twofold for the sodium sulphite treatment.     

Cell wall composition and degradability of forage stems following chemical and biological delignification

Chemical and biological delignification methods were used to investigate the relationship between the concentration and composition of lignin and degradation of forage cell walls. Stem material from lucerne (Medicago sativa L), smooth bromegrass (Bromus inermis Leyss) and maize (Zea mays L) stalks was treated with alkaline hydrogen peroxide, potassium permanganate, sodium chlorite, sodium hydroxide, nitrobenzene, and the lignolytic fungus Phanerochaete chrysosporium. Klason lignin and esterified and etherified phenolic acids were delermined. Cell wall neutral sugar and uronic acid composition and the extent of in-vitro degradability were measured. Chemical delignification generally removed lignin. but the fungal treatment resulted in the removal of more polysaccharide than lignin. The concentrations of esterfied and etherified p-coumaric and ferulic acids were generally reduced in treated cell walls; chlorite treatment preferentially removing p-coumaric acid whereas nitrobenzene treatment removed more ferulic acid. Syringyl moieties were completely removed from the core lignin polymer by nitrobenzene treatment of forage stems. Alkaline hydrogen peroxide and nitrobenzene were generally the most effective delignification treatments for improving polysaccharide degradability, with the grass species responding similarly to delignification whereas lucerne was somewhat less responsive. Fungal delignification, under these experimental conditions, did not improve cell wall degradability of these forages. Multiple regression and covariate analyses indicated that the lignin components measured were not powerful predictors of cell wall degradability. Neither the concentration nor the composition of the lignin fractions was consistently correlated with degradation. This lack of effect was attributed to the more generalised disruption of the cell wall matrix structure by delignification treatments.     

Degradation rate and chemical composition of different types of alkali-treated straws during rumen digestion

The degradation in the rumen and change in chemical composition of untreated and four samples of alkali-treated barley straw were studied in farm-scale treatments with ammonia, ‘dry’ sodium hydroxide and two different aqueous sodium hydroxide systems. The alkali treatments increased the fraction soluble in organic solvents and decreased the lignin content (determined as Klason lignin). The amounts of cellulose and arabinoxylans were not significantly changed. The degree of rumen degradability increased with the order of alkaline treatment given above. The degradation rates of dry matter and the NDF fraction were parallel to each other but varied considerably with the treatments. The two most efficient aqueous alkaline treatments showed a degradation rate four times that of untreated straw. For these two samples the arabinoxylans were solubilised faster than cellulose during the first part of the incubation. In the other samples the degradation of arabinoxylans and cellulose ran almost parallel. During the rumen incubation the lignin content of the straw residues increased for all samples. There was an inverse relationship between the content of p-coumaric and ferulic acids on the degree of digestibility. A relationship between acetyl groups and digestibility was not found.     

经胆碱氨基酸水溶液预处理的水稻秸秆中残余木质素对多糖酶水解抑制作用的研究

本文通过考察牛血清蛋白(BSA)的添加量和不同纤维素酶用量下其添加与否对水稻秸秆残渣中多糖酶水解效率的影响,以及比较不同木质素含量的底物对纤维素酶催化行为的影响来探讨[胆碱][氨基酸]处理水稻秸秆残渣后其残留木质素对纤维素酶活性的抑制作用大小。结果发现,低酶量下,BSA的添加对纤维素和半纤维素的酶水解降解度有轻微的促进作用,分别最多提高5%及7%;而高纤维素酶用量时,其促进作用甚微。且经该类离子液体处理后的不同木质素含量的水稻秸秆基本上对酶蛋白皆无显著吸附和抑制作用。可见,该类离子液体可去除部分木质素以提高酶分子对多糖底物的可及性,此为多糖酶水解效率提高的关键因素,而该法所形成的残余木质素对酶蛋白的非特异吸附或抑制作用相对较弱。     

Agriculture group symposium agriculture and pollution

The digestion of carbohydrates in straw and their substituent acetic and phenolic acids was studied using a slaughter technique. Two goats received non-alkali-treated and alkali-treated straws. The latter treatment increased the total apparent digestibility of cellulose by 56% and of hemicellulose by 110%. The ratio of cellulose to hemicellulose was altered in the rumen. For the non-alkali-treated straw the ratio decreased compared with the feed, whereas it increased for the alkali-treated straw. The ratio was kept almost constant post-rumen for both diets. The composition of the hemicellulose was also altered in the rumen compared with the straws fed. In particular the arabinose content decreased in both diets. Most of the polysaccharide digestion and absorption took place in the rumen. For the alkali-treated straw a hindgut fermentation could also be noted, especially for cellulose. Klason lignin was lost in the rumen but was recovered in the small intestine. Ferulic acid was more readily lost in the rumen than p-coumaric acid. Although acetyl groups were almost absent in the alkali-treated straw, they could still be found in the digesta of the animal. The slaughter technique appears to be a useful method when working with partitioning of digesta in ruminant animals.     

Biochemical characterization of wheat straw cell wall with special reference to bioactive profile

The core objective of the current study was to extract and characterize the wheat straw cell wall for its nutritional and bioactive profile. For the purpose, four different wheat straw varieties namely Ujala-16, Johar-16, Gold-16, and Galaxy-13 were procured from Ayub Agriculture Research Institute, Faisalabad, Pakistan. The whole research was conducted in three different phases. In first phase, nutritional composition and mineral profile of straw of different wheat varieties were determined. In second phase, wheat straw cell wall of different varieties was isolated and characterized for its important bioactive constituents, such as lignin, cellulose, hemicelluloses, phytosterol, and policosanol (PC) content. Results showed that straw of different wheat varieties contained 7.75–9.24, 3.98–5.06, 3.43–3.98, and 1.60–2.24 g/100 g moisture, ash, protein, and fat contents, respectively, whereas potassium, calcium, phosphorus, and magnesium were 1.19–2.03, 0.10–0.79, 0.10–0.98, 0.03–0.98 ppm, respectively. Moreover, lignocellulosic mass: cellulose 37.75–38.18 g/100 g raw material, lignin 15.67–16.07 g/100 g raw material, hemicelluloses 28.25–28.98 g/100 g raw material, was present in wheat straw and varied significantly among different varieties. In addition, phytosterol ranged from 854 to 1176 mg/kg in straw of different wheat cultivars, whilst PC from 196.09 to 236.48 mg/kg. Conclusively, wheat straw was an excellent source of many important bioactive moieties especially lignocelluloses and could have functional use.     

白腐菌对稻草秸秆中木质纤维素降解规律的研究

通过对傅里叶变换红外光谱(FTIR)谱图的分析和木质纤维素组分含量变化的测定,研究了3株白腐菌在50天培养期内降解稻草中木质纤维素的规律。结果表明,3株白腐菌对稻草秸秆中木质纤维素的降解具有一定的顺序和选择性,先降解半纤维素和木质素,再同时降解半纤维素、纤维素和木质素。从降解比例来看,白腐菌对半纤维素和木质素具有很好的降解优势,对半纤维素有较好的降解选择性。     

Depression of cellulose digestion by esterified cinnamic acids

Cinnamic acids commonly found in forages were esterified to cellulose and tested for their inhibition of cellulose degradation by mixed cultures of rumen microorganisms in vitro. Concentrations of cinnamic acids used in the cellulose esterification procedure were 20–100 g kg^?1, but measurement of actual ester concentrations by alkaline extraction were not possible for solka floc cellulose and ranged from 0–9.1 g kg^?1 for Whatman No. 54 filter paper. Cinnamic acid esters significantly depressed cellulose digestion. For both cellulose sources, caffeic acid was the most inhibitory compound. Solka floc cellulose digestion was also inhibited by sinapic acid, whereas p-coumaric and ferulic acids also depressed digestion of filter paper cellulose. The proportions of volatile fatty acids produced by fermentation of filter paper were not altered by cinnamic acid esters. Esterified cinnamic acids depressed cellulose digestion to a significantly greater extent than did free cinnamic acids. The data suggest that the natural form of cinnamic acids (ester-linked to cell wall fibre) are inhibitors of microbial digestion at concentrations reported to occur in forages.     

In-vitro digestibility of kale (Brassica oleracea) secondary xylem and parenchyma cell walls and their polysaccharide components

The cell walls of forage kale (Brassica oleracea L cvs Maris Kestrel, Vulcan and Thousandhead) stems were separated into parenchyma and secondary xylem fractions of high and low digestibility respectively. Previous work had shown that the lignified secondary xylem cell walls were the main components of the plant indigestible to ruminants. The cell walls were subjected to in-vitro digestion with fungal ‘cellulose’ and with rumen liquor. These two methods gave well correlated results except when secondary xylem cell walls were first delignified with chlorite, when their mean digestible organic matter in the dry matter (DOMD) was 41·9% with cellulase but 64·6% with rumen liquor. Chemical fractionation of the cellulase-digested residues showed that all fractions of the parenchyma cell walls, except the hemicelluloses from cv Thousandhead, were of high digestibility. In the secondary xylem cell walls the α-cellulose fraction had DOMD about 50%, the pectic fraction being more digestible and the hemicelluloses less digestible: but no one polysaccharide fraction in these cell walls was completely digested or untouched by digestion.     

Digestion of the dry matter, nitrogen, phosphorus, sulphur, calcium and detergent-fibre fractions of the seed and pod of Stylosanthes humilis contained in terylene bags in the bovine rumen

The rates of digestion of the separated seed and pods of Stylosanthes humilis in Terylene bags in the rumen of cattle were studied. The rates of disappearance of dry matter, N, P, S, Ca and detergent fibre fractions and lignin were examined. The magnitude of the errors of determination were assessed statistically for dry matter digestion. Four bag replications were necessary to detect differences of 20% between samples. The cell contents constituted 80% of the seed dry matter, but only 20% of the pod dry matter. Lignin content of seed was negligible (0.5%), but was high in pods (18%). The seed was rapidly digested (77% of dry matter after 48 h). This was largely due to almost complete digestion of the cell contents. The pod was only slightly digested (14% at 48 h). Protein and phosphorus appeared to be selectively solubilised, but calcium was only slowly solubilised in both seed and pod. The cell-wall constituents present were compared to the concentrations of hemicellulose and cellulose found in seed and pod by more definitive chemical methods.     

Effect of Barley Supplement on Microbial Fibrolytic Enzyme Activities and Cell Wall Degradation Rate in the Rumen

Three ruminally cannulated dry cows were used in a Latin square design to investigate the relationship between microbial fibrolytic enzyme activities and in sacco cell wall degradation of two gramineous hays, in which cell wall content ranged from 510 g kg⁻¹ DM for the regrowth to 687 g kg⁻¹ DM for the late harvested hay. Animals were fed twice daily a diet consisting of wheat straw, cocksfoot hay and ground barley in the ratios 10: 90: 0 (w/w), 10: 60: 30 (w/w) and 10: 30: 60 (w/w). For each diet and each hay, the in sacco degradation of cell wall polysaccharides and phenolic acids was determined. After 2 h and 23 h incubation time in the rumen, pH was measured in the bags, and fibrolytic activities (xylanase, avicelase, β-glycosidases) of the microbial population colonising hays in sacco were measured. Supplementation above 30% barley decreased the degradation rate of the cell wall polysaccharides, concomitantly with a decrease in polysaccharidase activities. The decrease in degradation rate was more marked for the regrowth than for the late harvested hay, for hemicelluloses than for cellulose and for ferulic than for p -coumaric acid. These differences did not appear to be related to microbial activities, which were similar between hays and between polysaccharidases, but rather to differences in accessibility of cell wall components to microbial enzymes, related to the composition of the forage and the cell wall architecture.     

Microbial degradation of normal maize and bm3 maize in the rumen observed by scanning electron microscopy

Samples of internodes and leaf blades from normal and bm3 maize (Zea mays L) harvested at dough to glazing stage were studied separately to determine their dry matter content, wall composition (NDF, ADF and ADL) and digestibility in sacco. For examination by light and scanning electron microscope, fragments 0·5 cm long were cut halfway along the internode beneath the female ear and on the corresponding blade. The wall and lignin contents of the bm3 maize were lower than in normal maize. The bm3 maize had a greater extent and faster rate of internode and blade disappearance in the rumen than normal maize samples. The histological structure of the two maizes was the same, but after 24 h in the rumen the parenchyma of the bm3 maize had degraded faster and the secondary walls of the fibres of its vascular bundles were degraded whereas those of normal maize had remained intact. After 72 h in the rumen the sclerenchyma of normal maize had changed little whereas that of the bm3 maize had much thinner walls and was abundantly colonised by rumen bacteria.     

纤维化学组分对再生植物纤维品质衰变影响的研究

通过选择性的脱除纤维细胞壁中的半纤维素和木素,研究半纤维素和木素含量对再生植物纤维品质衰变的影响.研究结果表明:随着半纤维素含量的减少,再生植物纤维的保水值降低,结晶度增加.当半纤维素含量从27.62%降至9.09%时,保水值下降了41%,纤维素结晶度从1.011增加到1.514.半纤维素含量与再生植物纤维成纸性能的变化有明显的线性相关性.较高含量的半纤维素可以缓解回用过程中再生植物纤维成纸性能的损失.同时发现木素对再生植物纤维性能的影响很小.木素含量与纤维的保水值、结晶度与成纸性能的变化没有明显的线性相关性.