Continuous single cell protein production fromCellulomonas sp. andCandida utilis grown in mixture on barley straw

Summary Continuous fermentations with mixed cultures of the cellulolytic bacteriaCellulomonas sp. and the yeastCandida utilis were examined. Fermentations were carried out in an aerated 5-l fermenter with different preparations of wet disintegrated barley straw as the cellulose source (3.6–4.2%). The straw was pretreated with NaOH (3.2–8.5 kg NaOH/100 kg dry straw) under high pressure and temperature in a feedstuff pellet press. The quantity of dry cell mass produced and the breakdown of the straw were measured. Crude protein and ash content in cell dry matter and residual fiber were determined. The experiments showed thatCellulomonas sp. andCandida utilis may be grown together in a continuous culture (dilution rate D=0.12–0.14 h^–1) for at least 3 days without washing out one of the organisms. Highest productivity was 1.39 g cell dry matter/l/h when using straw pretreated with 5.7% NaOH. The dry cell product contained 58–66% crude protein and up to 51% of the organic fiber dry matter was solubilized. The yield constants were 0.32–0.61 g cell dry matter per g solubilized organic fibers.     

The degradation and utilization of wheat-straw cell-wall monosaccharide components by defined ruminal cellulolytic bacteria

Cell walls (CW) of untreated wheat straw and sulphur-dioxide (SO₂)-treated wheat straw were used as model substrates for the hydrolysis and utilization of CW carbohydrates by pure cultures or pair-combinations of defined rumen bacterial strains. Fibrobacter succinogenes S85 and BL2 strains and their co-cultures with D1 were the best degraders of CW among ruminal cultures, solubilizing 37.2–39.6% of CW carbohydrates of untreated straw and 62.2–74.5% of SO₂-treated straw. Complementary action between Butyrivibrio fibrisolvens D1 and the F. succinogenes strains was identified with respect to co-culture growth and carbohydrate utilization. However, the extent of CW solubilization was determined mainly by the F. succinogenes strains. In both substrates, utilization of solubilized cellulose by F. succinogenes S85 and BL2 monocultures was higher than that of xylan and hemicellulose: 96.5–98.3%, 34.4–40.5% and 33.5–36.2%, respectively. Under scanning electron microscopy visualization, S85 and BL2 cells of the co-cultures comprised the most dense layer of bacterial cell mass attached to and colonized on straw stems and leaves, whereas D1 cells were always nearby. Stems and leaves of the untreated straw were less crowded by attached bacteria than that of the SO₂-treated straw. In both materials, the cell surface topography of S85 and BL2 bacteria attached to CW particles was specified by a coat of characteristic protuberant structures, polycellulosome complexes.     

Hydrolysate detoxification with activated charcoal for xylitol production by Candida guilliermondii

A detoxification method using activated charcoal with concentrated rice straw hemicellulosic hydrolysate improved the conversion of xylose to xylitol by the yeast Candida guilliermondii by 22%. This was achieved when the hydrolysate:charcoal ratio was 40 g g^–1, resulting in removal of 27% of phenolic compounds. Under this condition, the xylitol yield factor (0.72 g g^–1) and volumetric productivity (0.61 g l^–1 h^–1) were close to those attained in a semi-defined medium simulating hydrolysate sugars.     

Effect of inoculum level on xylitol production from rice straw hemicellulose hydrolysate byCandida guilliermondii

The effect of inoculum level on xylitol production byCandida guilliermondii was evaluated in a rice straw hemicellulose hydrolysate. High initial cell density did not show a positive effect in this bioconversion since increasing the initial cell density from 0.67 g L^–1 to 2.41 g L^–1 decreased both the rate of xylose utilization and xylitol accumulation. The maximum xylitol yield (0.71 g g^–1) and volumetric productivity (0.56 g L^–1 h^–1) were reached with an inoculum level of 0.9 g L^–1. These results show that under appropriate inoculum conditions rice straw hemicellulose hydrolysate can be converted into xylitol by the yeastC. guilliermondii with efficiency values as high as 77% of the theoretical maximum.     

Solid state fermentation and fractionation of oat straw by Basidiomycetes

Summary Seventee white-rot and brown-rot fungi were screened for their ability to fractionate the lignocellulose structure of oat straw through the preferential attack of lignin or cellulose. Fermentations were carried out under solid-state conditions with 25 g quantities of straw. The fermented straw was analyzed for weight loss, Klason lignin loss and cellulase digestion. All the fungi attacked both lignin and carbohydrate fractions causing 3–28% weight losses and 26–34 g/100 g enzymatic digestibility. Polyporus tulipiferae, Phanerochaete chrysosporium and Polyporus sp. were tested for the effects of various nitrogen, phosphate and carbon levels, incubation temperatures and incubation time. The three fungi had different responses to these factors.     

Statistical screening method for selection of important variables on xylitol biosynthesis from rice straw hydrolysate by Candida guilliermondii FTI 20037

Fourteen assays were conducted to study the influence of different variables, namely xylose concentration, inoculum level, agitation speed and nutrient supplementation, on xylitol biosynthesis by Candida guilliermondii FTI 20037. The maximum predicted values for xylitol yield (0.65 g g^–1) and xylitol productivity (0.66 g l^–1 h^–1) can be attained with rice straw hydrolysate containing 60 g xylose l^–1 without supplementation of ammonium sulfate, calcium chloride and rice bran extract, using 5 g inoculum l^–1, at 250 rpm. Xylose concentration and inoculum level were selected for further optimization studies.     

Effect of amendments,moisture and temperature on acetylene reduction in nile delta soils

Summary The pattern of N₂-ase activity in clay-loam soil of Nile Delta was determined. However, unamended soil showed somewhat low activity: an amount of 18–95 mg N₂ fixed/kg soil/year was calculated. Addition of glucose greatly enhanced such activity and efficiencies of N₂-fixation increased with decreasing carbon source concentration. Highest activities (800 n moles C₂H₄/gh^–1) and efficiencies (18.06 mg N₂/g glucose added) were reported in soil amended with 1% glucose, adjusted to 50% W.H.C. and incubated at 30°C. Enrichment of the soil with straw lead to a significant nitrogen gain particularly under water-logged conditions. During a short period of 16 days 5.8–9.3 mg N₂ were fixed/g straw added at the latter conditions.     

Three-year measurement of methane emission from an Indonesian paddy field

Yearly and seasonal (rainy and dry seasons) variations of CH₄ emission from a Sumatra paddy field were measured for 3 years. The mean CH₄ emission rates during the growth period were in the range of 16.0–26.1 mg CH₄ m^-2 h^-1 for the chemical fertilizer plots and 23.3–34.9 mg CH₄ m^-2 h^-1 for the plots with rice straw application, respectively. The increase in the amounts of CH₄ emission by rice straw application were from 1.3 to 1.6 times. There was no significant difference in the mean CH₄ emission rates between rainy and dry seasons.Total amounts of CH₄ emitted during the period of rice growth were in the ranges of 29.5–48.2 and 43.0–64.6 g CH₄ m^-2 for the plots applied with chemical fertilizer and those with rice straw application, respectively. Nearly the same amounts of CH₄ were emitted in the first and second half of the growth period, irrespective of rice straw application.     

Biocatalysts for production of chiral epoxides

Summary A number of bacterial strains, representing a range of genera, were isolated in pure culture with ethene or propene as the sole source of carbon and energy. The organisms included Aerococcus, Alcaligenes, Micrococcus and Staphylococcus spp. and a variety of Gram-negative, Gram-positive and Gram-variable mesophilic rods/ coccobacilli not yet identified. This suggests that the ability to utilize gaseous olefins is more widespread in nature than previously recognised. All 18 organisms tested stereospecifically formed R-1,2-epoxypropane (enantiomeric excess, ee=90–96%), R-1,2-epoxybutane (ee=90–98%) and trans-(2R,3R)-epoxybutane (ee=64–88%) from the corresponding olefins. In addition to Micrococcus sp. M90C, the substrate specificities of six other organisms were studied. The pattern of reactivity for the group of four ethene (M26, M90C, M93A, M186)- and two propene (M142, M156)-utilizers differed from that found with peracids, whereas the chemical reactivity of the substrate appeared to affect enzymatic epoxidations in Staphylococcus sp. M97B. Offprint requests to: M. Mahmoudian     

Degradation of a Lignin–Carbohydrate Substrate by Soil Fungi Producing Laccase and Cellobiose Dehydrogenase

The growth of nonsporulating mycelial fungi INBI 2-26(+), a producer of laccase; INBI 2-26(–), a producer of cellobiose dehydrogenase; and their mixed culture on lignin–carbohydrate substrates under conditions of submerged fermentation was studied. The degrees of degradation of lignin, cellulose, and hemicellulose of cut straw over 23 days amounted to 29.8, 51.4, and 72% for the laccase producer; 15.8, 33.9, and 59.1% for the cellobiose dehydrogenase producer; and 15.8, 39.4, and 64.5% for the mixed culture, respectively. The laccase activity in the medium when strain 2-26(+) was cultivated individually reached its maximum on day 28; the activity of cellobiose dehydrogenase of strain 2-26(–), on days 14–28. A method for determining cellobiose dehydrogenase activity in the presence of laccase was developed. In the mixed culture, both enzymes were formed; however, the level of laccase synthesis was 1.5-fold lower compared to that of strain 2-26(+), while synthesis of cellobiose dehydrogenase was similar to that of the corresponding producer. Cellobiose dehydrogenase failed to boost the action of laccase while degrading the lignin of straw.     

Biotreatment of nitrate-rich industrial effluent by suspended bacterial growth

Of the 29 potentially denitrifying organisms isolated from a denitrifying reactor (DNR) of a fertilizer company, two isolates; I-4 and I-5 were recognized as denitrifiers. Under aerobic conditions, with fusel oil as the carbon source, the organisms decreased nitrate from 1200 mg l^–1 to 100 mg l^–1 in 48 h. Optimal growth conditions for biological removal of nitrate were established in batch culture. The system was scaled up to 4-L and 50-L bioreactors under continuous culture conditions. Up to 95–100% nitrate removal was achieved in the 50-L bioreactor at a COD:NO₃–N ratio of 3.45 with a retention time of 48 h. The isolates showed 1.5 fold higher denitrifying activity than reported previously.     

Production of extracellular enzymes during the solubilisation of straw by Thermomonospora fusca BD25

The production of three extracellular enzymes during the solubilisation of ball-milled wheat straw by seven actinomycete strains, was examined. A general correlation was observed between the production of extracellular enzymes (xylanases, endoglucanases and peroxidases) and the formation of the solubilised lignocellulose intermediate product (APPL), with the thermophilic actinomycete Thermomonospora fusca BD25 exhibiting greatest extracellular enzyme activity and highest APPL production. Production of all three enzymes; endoxylanase, endoglucanase and peroxidase, and lignocellulose solubilisation, occured during primary growth with maximum activity at the end of the exponential phase (48–96 h). The inducibility and stability of extracellular enzymes from T. fusca were further characterised. When xylan replaced ball-milled wheat straw as the growth substrate, reduced enzyme activities were observed (28–96% reduction in enzyme activities), whereas carboxymethylcellulose was found to be a poor inducer of all three enzyme activities (80–100% reduction in enzyme activities). The pH and temperature optima for extracellular enzyme activities from T. fusca was found to be pH 7.0–8.0 and 60°C, respectively. Analysis of concentrated crude supernatant from T. fusca by native polyacrylamide gel electrophoresis revealed the existence of two non-haem peroxidases. The stability of the extracellular lignocellulose-degrading enzymes for T. fusca suggest their suitability for future biotechnological processes such as biobleaching.     

Farmland treatment and utilization of straw pulp mill effluents

Straw pulp production accounts for 74% of the total raw pulp production in China. The pulping waste, containing high contents of silicate and high pH, is difficult to treat. Instead of the conventional alkaline process, ammonia sulfite pulping discharges neutral effluent with nitrogen, phosphorus, potassium, sulfur and organic matter which are potential fertilizers and water resources for agriculture. The effect of effluent from ammonia sulfite pulping on rice growth and yield, was studied in Baigezhuang farm, Hebei province. Diluted rice straw digester waste or composite waste were irrigated on a paddy field. In plot tests, rice output increased by 16.2–25.3% with 0.7–6% pulp digester waste and by 22.9–37.9% with 2.7–5.4% composite waste. In field tests, rice output increased by 8.6–15.9% with 2.7–4.1% composite waste. Due to clean water dilution and paddy purification, the removal efficiency of biochemical oxygen demand (BOD), chemical oxygen demand (COD), suspended solid (SS), sulfide and phenol can reach over 97%. Rice quality also tested had higher protein content than the control. The study suggests that the effluents from ammonia sulfite pulping can be widely used in the farming industry. The ammonia sulfite process is a good approach to overcome pulp pollution in rural areas.     

Kinetic study of hydrolysis of xylan and agricultural wastes with hot liquid water

We investigated the kinetics of hot liquid water (HLW) hydrolysis over a 60-min period using a self-designed setup. The reaction was performed within the range 160–220 °C, under reaction conditions of 4.0 MPa, a 1:20 solid:liquid ratio (g/mL), at 500 rpm stirring speed. Xylan was chosen as a model compound for hemicelluloses, and two kinds of agricultural wastes–rice straw and palm shell–were used as typical feedstocks representative of herbaceous and woody biomasses, respectively. The hydrolysis reactions for the three kinds of materials followed a first-order sequential kinetic model, and the hydrolysis activation energies were 65.58 kJ/mol for xylan, 68.76 kJ/mol for rice straw, and 95.19 kJ/mol for palm shell. The activation energies of sugar degradation were 147.21 kJ/mol for xylan, 47.08 kJ/mol for rice straw and 79.74 kJ/mol for palm shell. These differences may be due to differences in the composition and construction of the three kinds of materials. In order to reduce the decomposition of sugars, the hydrolysis time of biomasses such as rice straw and palm shell should be strictly controlled.     

Effects of Wheat Straw Incorporation on the Availability of Soil Nutrients and Enzyme Activities in Semiarid Areas

Soil infertility is the main barrier to dryland agricultural production in China. To provide a basis for the establishment of a soil amelioration technical system for rainfed fields in the semiarid area of northwest China, we conducted a four—year (2007–2011) field experiment to determine the effects of wheat straw incorporation on the arid soil nutrient levels of cropland cultivated with winter wheat after different straw incorporation levels. Three wheat straw incorporation levels were tested (H: 9000 kg hm^-2, M: 6000 kg hm^-2, and L: 3000 kg hm^-2) and no straw incorporation was used as the control (CK). The levels of soil nutrients, soil organic carbon (SOC), soil labile organic carbon (LOC), and enzyme activities were analyzed each year after the wheat harvest. After straw incorporation for four years, the results showed that variable straw amounts had different effects on the soil fertility indices, where treatment H had the greatest effect. Compared with CK, the average soil available N, available P, available K, SOC, and LOC levels were higher in the 0–40 cm soil layers after straw incorporation treatments, i.e., 9.1–30.5%, 9.8–69.5%, 10.3–27.3%, 0.7–23.4%, and 44.4–49.4% higher, respectively. On average, the urease, phosphatase, and invertase levels in the 0–40 cm soil layers were 24.4–31.3%, 9.9–36.4%, and 42.9–65.3% higher, respectively. Higher yields coupled with higher nutrient contents were achieved with H, M and L compared with CK, where these treatments increased the crop yields by 26.75%, 21.51%, and 7.15%, respectively.     

Nitrogen transformation in soil during humification of straw labelled with15N

Summary The decomposition and humification of oat straw labelled with¹⁵N were followed in the soil during 80 days. The influence of NaNO₃ and (NH₄)₂ SO₄ on these processes were also investigated. It was ascertained that addition of NH₄–N acted more efficiently than NO₃–N on both the decomposition of straw and the mineralization of the organic nitrogen compounds of the soil. In the presence of NH₄–N, straw¹⁵N predominated in humic acids, while in the presence of NO₃–N it predominated in fulvic acids.The incorporation of straw¹⁵N into the humic compounds occurred in proportion to the progressing decomposition of straw. The greatest similarity in the proportions of soil-N and straw-¹⁵N in isolated fractions was ascertained after 80 days of incubation in the presence of NH₄–N.     

Microwave-assisted conversion of lignocellulosic biomass into furans in ionic liquid

Production of 5-hydroxymethylfurfural (HMF) and furfural from lignocellulosic biomass was studied in ionic liquid in the presence of CrCl₃ under microwave irradiation. Corn stalk, rice straw and pine wood treated under typical reaction conditions produced HMF and furfural in yields of 45–52% and 23–31%, respectively, within 3 min. This method should be valuable to facilitate energy-efficient and cost-effective conversion of biomass into biofuels and platform chemicals.     

An electron microscopic study of picoplanktonic organisms from a Small Lake

Picoplankton, both prokaryotic and eukaryotic, are distinguished from other aquatic organisms by their small size (0.1–2.0 m). Such organisms were recovered from waters of a small oligotrophic lake using screens, filters, and high-speed centrifugation. The majority of the picoplankton were unable to form visible colonies on common media. Cells examined in thin sections by electron microscopy showed that 60–75% of the cells had an average diameter after dehydration of 0.48–0.51 m. The maximum dimensions of the rest of the cells ranged from 0.56–1.81 m. Using details of ultrastructure, cells were classified as prokaryotic or eukaryotic. Phototrophs present included two cyanobacterial morphotypes (5–6%) and two eukaryotic algae (less than I%). The arrays of intracytoplasmic membranes in 18–20% of the cells were suggestive of methanotrophic rods and chemoautotrophs. Relatively few prosthecate bacteria were observed in the water column samples. The smallest cells (1–2%) contained magnetosomes, the presence of which were confirmed by x-ray spectroscopy. Iron was also detected in the envelopes of some rod shaped cells by the same technique. The study of in situ picoplankton populations using TEM coupled with other techniques may provide better understanding of picoplankton biomass.     

Integrating sub-lethal heating with Brassica amendments and summer irrigation for control of Macrophomina phaseolina

Effects of varying intensities of sub-lethal heating were ascertained in improving the efficiency of Brassica amendments and summer irrigation on survival of Macrophomina phaseolina, a dry root rot pathogen. Sub-lethal heating (45–55°C) of M. phaseolina infested dry soil reduced the viable propagules by only 12.8% in a period of 90 days. One summer irrigation without sub-lethal heating caused 33.9% reduction in M. phaseolinapropagules, which improved to 43.3% when it was combined with 60 days of sub-lethal heating. Addition of the Brassicaamendments to irrigated soil resulted in significant reduction (60.4–71.6%) in counts of M. phaseolinabut this reduction improved (89.4–96.1%) when sub-lethal heating was combined with amendments. Mustard oil-cake (0.18% w/w) was found to be the most effective with reduction but a 94% inoculum reduction by mustard pod straw (0.36% w/w) was also achieved at 0–30 cm soil depth under similar conditions. Moderate heat level could not exert detectable weakening effect on M. phaseolinapropagules. These results suggest a practical cultural control of soil-borne pathogens by combining sub-lethal heating, Brassica amendments with one summer irrigation.     

On the concentration of acetic acid in straw and soil

Summary Freshly harvested wheat straw contained 0.096 g water g^–1 dry straw and 180 mM acetic acid. The straw absorbed water more rapidly from wet soil. The concentration of acetic acid fell to about 10 mM within 6 h of incorporation of straw in the soil and then remained relatively constant for a period of 12 days, irrespective of soil moisture content. In soil at its maximum water holding capacity after gravitational drainage, the decline in acetic acid concentration (c) with distance (d) from wheat or barley straw was exponential, with c=c_o e^–nd where c_o is the concentration of acetic acid at the straw surface and n is a constant (0.46 for barley and 0.42 for wheat straw). The presence of acetic acid seems to be a major cause of poor establishment and growth when seeds and seedling roots come into contact with straw.