Agar quality of commercial agarophytes from different geographical origins: 1. Physical and rheological properties

Six economically important species ofGracilaria, from a number of commercial sources around the world, andGracilariopsis lemaneiformis, collected from two Japanese localities, were used as the sources of raw material for the evaluation of agar quality. Agar-agar was extracted by pretreatment with various concentratrions of NaOH (0%, 3%, 5%, 7%, 10%) incubated at 80 °C for 2 h. Agar yield, viscosity, dynamic gelling and melting temperature and gel texture were determined for 1.5% agar gels. The highest agar yield was obtained fromG. gracilis from Argentina (39.5%), while the lowest was from BrazilianG. gracilis (13.37%). Dynamic gelling temperature was highest in the agar fromG. gracilis from Turkey (59 °C) and lowest in the non-alkali treated agar isolated fromG. edulis from Indonesia (46 °C). Melting temperature ranged from 96 °C in the agars from the JapaneseGracilariopsis andG. chilensis from Chile to 69 °C in the non-alkali treated agar fromG. edulis from Indonesia. In general, all species produced an agar with high gel strength after treatment with 5% NaOH, except forG. chilensis and the twoGracilariopsis species, which produced an agar with high gel strength after treatment with 3, 7 and 10% NaOH. The highest gel strength (2056 ± 13.6 cm^–2) and hardest gel (261 ± 19.89 g mm^–2) were obtained fromG. lemaneiformis from Japan (Oita Prefecture) after treatment with 7 and 10% NaOH respectively. The lowest gel strength (351 ± 93 cm^–2) was obtained fromG. gracilis from Brazil after treatment with 3% NaOH. The softest gel (66.31 ± 9.63 g mm^–2) was isolated fromG. tenuistipitata from China, after treatment with 3% NaOH. The most flexible gel (11.62 ± 0.31 g mm^–2 × 10²) was obtained fromG. chilensis from Chile after treatment with 3% NaOH.Author for correspondence     

Alkali treatment and fermentation of straw for animal feed

The feed value of annual ryegrass straw was improved by treatment with various concentrations of NaOH or NH₃ followed by fermentation of the treated straw with a mixed culture of Cellulomonas sp. and Alcaligenes faecalis. Laboratory feeding trials with voles showed that NaOH or NH₃ treatment considerably increased the feed efficiency of straw, but apparently gave a poorly palatable product. Fermentation tended to decrease the in vitro rumen digestibility (IVRD) of alkali-treated straw. The fermentations were carried out aerobically on a semisolid straw matrix having 11–86% moisture. Treatment by both NaOH and NH₃ increased the IVRD of straw. NH₃ also increased the nitrogen content in straw. The optimum condition for alkaline treatment of the straw was 4–6% NaOH for 1 hr or with 3% NH₃ for four weeks at room temperature. A minimum of 63% moisture was needed for significant fermentation of the straw. The combined effects of NaOH treatment and fermentation more than doubled crude protein, doubled crude fat, and increased IVRD by 75%. The NH₃ plus fermentation treatment tripled crude protein, doubled crude fat, and increased IVRD by 60%. Acetic acid was the main volatile fatty acid in the fermented straw.     

Structural and physico-chemical characterization of hemicelluloses from ultrasound-assisted extractions of partially delignified fast-growing poplar wood through organic solvent and alkaline solutions

One organic and three alkaline hemicellulosic fractions were isolated by an ultrasound-assisted extraction which partially delignified the fast-growing poplar wood. Successive treatments were conducted with dimethyl sulfoxide under ultrasonic irradiation at 570 W, 25 °C for 30 min, 70% ethanol containing 1% NaOH, 3% NaOH and 6% NaOH at 75 °C for 3 h, respectively. The four hemicellulosic fractions obtained were comparatively studied by sugar analysis, alkaline nitrobenzene oxidation of bound lignin, GPC, FT-IR, 1D and 2D NMR spectroscopy as well as TGA and DTA. The results showed that the ultrasonic treatment and sequential extractions with three different concentrations of NaOH led to a release of 75.5% of the original hemicelluloses and 96.2% of the lignin. All four purified hemicellulose fractions contained relatively low amounts of associated lignin, ranging between 0.96 and 3.10%. In addition, the hemicellulosic fraction H₄ isolated with 6% NaOH is formed by a linear backbone of four (β-1 → 4)-xylopyranosyl residues and at least one of the xylose residues is monosubstituted at C-2 by a 4-O-methylglucuronic acid, giving a typical ratio of 4-O-methyl glucuronic acid to Xyl of 1 to 4.     

Extraction of chitin and chitosan from housefly,Musca domestica,pupa shells

Chitins and chitosans are some of the most abundant natural polysaccharide materials, and are used to increase innate immune response and disease resistance in humans and animals. In this work, chitin and chitosan from housefly, Musca domestica, pupa shells were obtained by treatment with HCl and NaOH. For chitin extraction, 2 N HCl and 1.25 N NaOH solutions were used to achieve decalcification and deproteinization, respectively. For chitosan extraction, 50% NaOH solution was used to achieve deacetylation. The yields of chitin and chitosan from pupa shells of M. domestica were 8.02% and 5.87%, respectively. The deacetylations of chitosan (from chitin C1 and C2) were 89.76% and 92.39%, respectively, after the first alkali treatment with 50% NaOH (w/w) solution at 105 °C for 3 h and 5 h, respectively. The viscosities of the chitosans (from chitin C1 and C2) were 33.6 and 19.2 cP, respectively.     

Optimization of ultrasonic-assisted extraction process of Poria cocos polysaccharides by response surface methodology

Polysaccharides production from Poria cocos was carried out using aqueous NaOH with the assistance of ultrasonic. Experimental design was used to investigate the effect of three parameters (extraction time, extraction concentration of NaOH, and ratio of aqueous NaOH to raw material) on polysaccharides yields. The ranges of the factors investigated were 1–3 min for extraction time (X₁), 0.5–1.0 mol/L for extraction concentration of NaOH (X₂), and 30–50 for ratio of aqueous NaOH to raw material (X₃). The statistical analysis of the experiment indicated that extraction concentration of NaOH had significant effect on P. cocos polysaccharides yields. The central composite design showed that polynomial regression models were in good agreement with the experimental results with the coefficients of determination of 0.9935 for P. cocos polysaccharides yield. The optimal condition for P. cocos polysaccharides yield within the experimental range of the variables studied was at 2.44 min, 0.789 mol/L, and 53.0. At this condition, the predicted yield of polysaccharides extracted was 82.3%.     

Effect of level of sodium hydroxide treatment and volume of solution on the nutritive value of maize cobs

Maize cobs were chosen as a test material as these are produced centrally in home-steads during shelling and therefore collection and transport costs are eliminated. The knowledge gained from the improvement of their nutritive value by the dry NaOH treatment method can be adapted easily.The effect of treatment rates was tested with rams in three experiments. In the first experiment, coarsely ground maize cobs were treated at 1.67, 3.33 and 5.0 kg NaOH/100 kg cobs dry matter (DM) and formed 52% of the rations on an air-dry basis. Digestibility increased significantly (P < 0.05) and a regression equation y = 51.57 + 2.28x was calculated in which y = DM digestibility, and x = kg NaOH/100 cobs DM. In the second experiment, maize cobs treated at 2.5, 5.0, 7.5 and 10.0 kg NaOH/100 kg cobs DM formed 67% of the ration and the digestibility of DM, organic matter (OM), cell wall constituents (CWC) and crude fibre (CF) were markedly increased from 2.5 to 5.0 kg NaOH treatment levels and no further responses were obtained thereafter. In the third experiment, maize cobs formed 61% of the rations and the material was treated at 0, 2.5, 5.0 and 7.5 kg NaOH. Digestibility of DM, OM, CF and energy was significantly (P < 0.05) increased at 5.0 and 7.5 kg NaOH levels. Voluntary feed intake followed the same trend, values for g DM intake per kgW^0.75 per day being 60.5, 83.3, 95.7 and 96.0 in the order of increasing NaOH treatment rates. The total digestibility DM, OM and energy intakes were more closely correlated to voluntary feed intake than to digestibility coefficients. It is concluded that 5 kg NaOH/100 kg DM is optimum for both digestibility and voluntary feed intake.The volume of NaOH solution was varied from 25 to 200 l per 100 kg DM of maize cobs in four experiments and the digestibility of DM, OM and CWC with sheep was not affected by this factor, except that the smallest volume (25 l) depressed digestion slightly, apparently because of the difficulty of getting effective mixing with small volumes. It is concluded that 50 l per 100 kg DM of maize cobs is the minimum volume.     

Evaluation of chemical pretreatment for enzymatic solubilization of rice straw

Summary Rice straw was treated with NaOH, peracetic acid (PA), and sodium chlorite (NaClO₂). Quantitative changes in the composition of the treated straw, crystallinity of the treated straw and extracted cellulose, and susceptibility of the treated straw to Trichoderma reesei cellulase were studied. The alkali treatment resulted in a remarkable decrease in hemicellulose as well as lignin. Consequently, the recovery of residual straw after NaOH treatment was lowest among the three chemical reagents evaluated. The treatment with PA or NaCIO₂ resulted in a slight loss in hemicellulose and cellulose in the straw. The three chemical treatments caused little or no breakdown of the crystalline structure of cellulose in the straw. The treated straw was solubilized with the culture filtrate of T. reesei. The degree of enzymatic solubilization relative to the amount of residual straw was 69% after treatment with 0.25 N NaOH, 42% after treatment with 20% PA, and 50% after treatments with NaClO₂ (twice). The degree of enzymatic solubilization relative to the amount of the untreated straw, however, was 30% after treatment with 0.25 N NaOH, 32% after treatment with 20% PA, and 37% after treatments with NaClO₂ (twice).     

Evaluation of different alkali treatments of bagasse pith for cultivation of Cellulomonas sp.

Biomass production of Cellulomonas was optimal with 1% (w/v) bagasse pith pre-treated with either 0.2 M NaOH for 1 h at 80°C or 0.4 M NaOH for 40 h at 28 to 30°C. Growth was similar to that obtained with a more severe treatment used as control and compared well with other reports for cellulolytic bacteria cultivated on pre-treated bagasse pith.     

Single cell protein production from bagasse pith pretreated with sodium hydroxide at room temperature

Summary Previous publications have revealed that a pretreatment of lignocellulosic wastes is necessary if they are to be employed as the hydrocarbon source of single cell protein production. A hot alkaline treatment is the most common.We have treated sugar cane bagasse pith with 1% NaOH solution at room temperature, at a NaOH/pith ratio of 10%. Different contact times were used in the experiments. The shortest contact period required for maximum protein production was 24 h at 25° C. A mixed culture of Cellulomonas sp. and Bacillus subtilis was used in the experiments. The values obtained for hemicellulose and cellulose in the treated pith did not differ greatly from those of untreated pith, in contrast the amount of lignin was 33% lower in the treated pith. The effect of reutilization of the alkaline liquor used for the pretreatment of pith upon protein production was also investigated. With four recyclings, there was a NaOH saving of 34.4 kg per 100 kg produced protein as compared to when the liquor was only used once.The quality of the resulting effluents, as measured by the chemical oxygen demand (COD), proved to be very similar for both types of treatment.     

Production of chitin and chitosan from the exoskeleton of adult two‐spotted field crickets (Gryllus bimaculatus)

Chitin and chitosan were extracted from all specimens of Type I and II two‐spotted field crickets (Gryllus bimaculatus) following chemical treatment with an acid and alkali. For chitin extraction, 2 N HCl and 1.25 N NaOH solutions were used to achieve demineralization and deproteinization, respectively. For chitosan extraction, 50 % NaOH (w/v) and 50 % NaOH (w/w) solutions were used to achieve deacetylation. Chitosan yielded from adult exoskeletons of G. bimaculatus in Test A of Type I was 1.76 and 8.40 % on a fresh weight (FW) and dry weight (DW) basis, respectively, after treatment with 50 % NaOH (w/v) at 95°C for 3 h. Furthermore, the chitosan yielded in Test D of Type II was 1.79 and 7.06 % on FW and DW basis, respectively, after treatment with 50 % NaOH (w/w) at 105°C for 3 h. The average yield of chitin and chitosan was 2.42 and 1.65 % on a FW basis, and 10.91 and 7.50 % on DW basis, respectively. The deacetylation (%) of chitosan extracted from adult exoskeletons in Tests A, B, C1, C2, D1, and D2 were 81.2 %, 14.5 %, 19.6 %, 90.7 %, 17.1 %, and 95.5 %, respectively. The viscosities of the chitosans extracted from adult exoskeletons in Tests A, C2, and D2 were 32.0, 21.6, and 62.4 cP (centi Poise), respectively. The molecular weight of chitosan from adult exoskeletons of G. bimaculatus was 308.3 kDa. Our results indicate that adult exoskeletons of G. bimaculatus could be used as a source of chitin and chitosan for use as functional additives in industrial animal feeds.     

Enzymatic Saccharification of Pretreated Wheat Straw by T. Reesei Cellulases and A. Niger β-Glucosidase

Three methods of wheat straw treatment (with NaOH, H₂O₂ and butylamine) and its subsequent saccharification by Trichoderma reesei cellulases and Aspergillus niger β-glucosidase are reported. The treatment of straw with NaOH for 1 h in the autoclave (120°C) caused a great loss of the hemicellulose content and a partial removal of lignin, provoking an increase of the cellulose content (from 24% to 69%) in the residue. When the straw was pre-treated with H₂O₂ at 25°C for 20 h, the relative content of cellulose in the straw increased (from 24% to 52%) due to the solubilisation of hemicellulose.

The effect of varying the hydrolysis parameters (enzyme and substrate concentration, temperature and pH) was studied in order to maximise the yield of sugars. Under the best conditions and after 48 h with a mixture of 2:1 (w/w) cellulase/β-glucosidase (with a concentration of 7 and 0.1 U ml^-1, respectively) the native, NaOH-treated and H₂O₂-treated straw material were degraded to reducing sugars for 28%, 89% and 97% respectively.     

CELL DIVISION AND THE ROLE OF THE PRIMARY WALL IN THE FILAMENTOUS DESMID ONYCHONEMA LAEVE (CHLOROPHYTA)1

Cell division and the role of the primary wall in filament formation in the desmid Onychonema laeve Nordst. were investigated by transmission and scanning electron microscopy. In addition, sequential chemical extractions and enzyme treatments were performed, on cell walls of intact filaments. Interphase cells are deeply constricted, consisting of two semicells, each elliptical in front view and circular in side view. In addition to two short lateral spines, each semicell has two apical processes that originate on opposite sides at an angle of about 15° from the central axis and overlap the adjacent cell. Division is initiated as in other desmids by a slight separation of the semicells and development of a girdle of primary wall material at the isthmus. In O. laeve the girdle of primary wall expands to form a spherical vesicle (termed a division vesicle) between the separating semicells. Nuclear division and septum formation occur in this vesicle when it is nearly the full diameter of the filament. Morphogenesis of the apical processes begins with completion of the septum, before the secondary wall appears. At maturity each apical process is surrounded by a thick layer of both secondary and primary wall, except that its capitate tip protrudes through the shroud of primary wall. Sequential treatment with hot ammonium oxalate, 4% NaOH, 17.5% NaOH and 10% chromic acid or various enzyme solutions did not cause filament breakage. SEM and TEM views of O. laeve after these treatments show intact secondary walls and intact primary wall material covering and connecting the apical processes of adjacent cells. It is the persistence of the primary wall between cells and around the apical processes that maintains the long, unbranched filamentous morphology of Onychonema laeve.     

Biocontrol of Botrytis cinerea in apple fruit by Cryptococcus laurentii and indole-3-acetic acid

This study evaluated the effect of a yeast antagonist Cryptococcus laurentii and a plant regulator indole-3-acetic acid (IAA) on inhibition of Botrytis cinerea infection in harvested apple fruit. The results showed that the combined treatment with C. laurentii and IAA at 20 μg/ml was a more effective approach to reduce the gray mold rot in apple wounds than the C. laurentii alone. After 4 days of incubation, gray mold incidence in the combined treatment with C. laurentii and IAA was about 18%, which was a 50% reduction in incidence compared to the treatment with C. laurentii alone. Although IAA had no direct antifungal activity against B. cinerea infection when the time interval between IAA treatment and pathogen inoculation was within 2 h, application of IAA strongly reduced gray mold infection when IAA was applied 24 h prior to inoculation with B. cinerea in apple fruit wounds. Moreover, combination of IAA and C. laurentii stimulated the activities of superoxide dismutase, catalase and peroxidase with above 1.5-fold higher than that treatment with C. laurentii alone at 48 h. Therefore, combination of C. laurentii with IAA, which integrated the dual biological activity from the antagonistic yeast and plant regulator, might be developed to be a useful approach to control gray mold in harvested apple fruit.     

Biological control by Clonostachys rosea as a key component in the integrated management of strawberry gray mold

Gray mold, caused by Botrytis cinerea, is an important strawberry disease. As gray mold control is difficult, there is a need to evaluate integrated methods to successfully manage the disease. The efficiency of integrating Clonostachys rosea sprays, fungicide sprays, and crop debris removal to manage gray mold was evaluated in field experiments conducted in 2006 and 2007. Leaf colonization by C. rosea (LAC), average number of B. cinerea conidiophores (ANC), gray mold incidence in both flowers (Iflower) and fruits (Ifruit), and yield were evaluated weekly. In both years, LAC was higher in the treatments with no fungicide. When compared to the check, ANC, Iflower and Ifruit were most reduced in treatments that included C. rosea sprays. Maximal reductions were achieved with the combination of C. rosea sprays, fungicide sprays and debris removal (96.62%, 86.54% and 65.33% reductions of ANC, Iflower and Ifruit, respectively). Otherwise, maximal yield (103.14% increase as compared to the check) was achieved with the combination of the three treatments. With just C. rosea sprays, ANC, Iflower and Ifruit were reduced by 92.01%, 68.48% and 65.33%, respectively, whereas yield was increased by 75.15%. Considering the individual effects, application of C. rosea was the most efficient treatment. Chemical control was effective only in plots without debris removal. Elimination of crop debris was the least effective method in reducing gray mold incidence in both flowers and fruits. The integrated control approach enhanced the efficacy of the individual methods of gray mold control and provided high strawberry yield. An important component of this integrated approach it the biological control with C. rosea.     

Extraction of hemicellulose from ryegrass straw for the production of glucose isomerase and use of the resulting straw residue for animal feed

The hemicellulose fraction of ryegrass straw was extracted with NaOH and used for the production of glucose isomerase by Streptomyces flavogriseus. The level of hemicellulose extracted increased proportionately with increasing NaOH concentration up to about 4%, then the rate of increase slowed down. Hemicellulose extraction was facilitated by the combined application of heat and NaOH. Approximately 15% hemicellulose (12% as pentosan) could be obtained by treating straw with 4% NaOH for either 3 hr at 90°C or 24 hr at room temperature. The highest level (3.04 units/ml culture) of intracellular glucose isomerase was obtained when the organism was grown at 30°C for two days on 2% straw hemicellulose. The organism also produced a high yield of glucose isomerase on xylose or xylan. The NaOH-treated straw residue, after removal of hemicellulose, had approximately 75% higher digestibility and 20% higher feed efficiency for weanling meadow voles than untreated straw. Thus, the residue could be used as animal feed. A process for the production of glucose isomerase and animal feed from ryegrass straw was also proposed.     

次氯酸钠离析法观测甘蓝叶片脉序的处理条件优化筛选

次氯酸钠(NaClO)离析法主要用于植物叶片表皮的观测, 在研究过程中发现该法也可用于叶片脉序的观测。以甘蓝(Brassica oleracea var. capitata)为实验材料, 采用二次正交设计方法对水煮时间、NaOH浓度、NaOH处理时间、NaClO浓度、NaClO处理温度和处理时间等各种处理条件进行优化筛选, 以期得到适合于甘蓝叶片脉序观测的最佳处理条件组合。实验结果表明, 新鲜甘蓝叶片水煮3分钟, 10%的NaOH溶液60°C水浴处理2.5小时, 3%的NaClO溶液40°C水浴离析2小时,叶片脉序的观测效果最佳。     

Evidence for Covalently Attached p-Coumaric Acid and Ferulic Acid in Cutins and Suberins

p-Coumaric acid (4-hydroxycinnamic acid) and ferulic acid (4-hydroxy-3-methoxycinnamic acid) have been identified as constituents of cutin. Their reduction products were isolated from a phenolic fraction released from the cutin of the fruits of apple, peach, pear, and two varieties of tomato and apple leaf by treatment with LiAlH(4) or LiAlD(4). They were identified by combined gas chromatography and mass spectrometry. p-Coumaric acid was present in all samples of cutin (0.07-0.53% by weight), whereas only peach and pear cutin contained measurable amounts of ferulic acid (0.007% and 0.035%, respectively). Both p-coumaric acid and ferulic acid were identified to be constituents of the insoluble material recovered after partial hydrolysis (12-42% loss) of cutin in 1 m NaOH at 80 C. A significant part (48%) of the p-coumaric acid contained in tomato cutin was contained in the insoluble material recovered after partial degradation (7.4%) of this cutin with 0.01 m NaOH. These data indicate that these phenolic components are tightly (possibly covalently) bound to cutin. Similar analysis of the phenolic fractions from the suberins of potato, sweet potato, turnip, rutabaga, carrot, and red beet revealed that they contained only ferulic acid (0.05-0.22%). Ferulic acid was identified as a constituent of the insoluble material recovered after partial hydrolysis of potato and beet suberins (34% and 32% loss, respectively) in 1 m NaOH at 80 C. A major part (65%) of the ferulic acid contained in potato suberin was contained in the insoluble material recovered after partial (26.8% loss) degradation of this suberin with 0.01 m NaOH. Ferulic acid appears to be tightly (probably covalently) bound to suberin.     

Optimization of the yield and quality of agar from Gracilariopsis lemaneiformis (Gracilariales) from the Gulf of California using an alkaline treatment

The effect of several alkali treatments on the yield, gel strength, rheology, and chemical characteristics (quality) of the agar obtained from Gracilariopsis lemaneiformis from the Gulf of California was analyzed using different alkali concentrations, temperatures and treatment times. In the first stage of the experiment, all treatments lasted 60 min and the NaOH concentrations (2.5, 3.0, 4.0, 5.0, 6.0%) and temperature (80, 90, 100°C) varied. At constant time, temperature played the predominant role, promoting an increase in agar gel strength. Based on the best treatment conditions found (4% and 5% NaOH, and 90°C and 100°C temperature), in the second stage different treatment times (15, 30, 60, 90, 120 min) were used. Since agar yields were not significantly different among temperatures and times, the optimal conditions to obtain best quality agar were those providing the highest gel strength. Treatment time played an important role in increasing gel strength. Maximum gel strength (Nikan, 954 g cm⁻²) was obtained with 5% NaOH at 100°C after 90 min of treatment, though these conditions resulted in an agar yield reduction of 25.5% relative to native agar. This treatment proved to efficiently yield G. lemaneiformis agar that will meet the commercial quality requirements regarding gel strength, 3,6 anhydrogalactose and sulfate content, as well as rheology and hysteresis. Enrique Hernández-Garibay holds a CONACyT scholarship.     

Alkali-induced conversion of beta-chitin to alpha-chitin

Crystal conversion of beta-chitin to alpha-chitin by aq. NaOH treatment was studied for a highly crystalline beta-chitin sample from diatom spine. The minimum NaOH concentration to cause swelling was between 25% and 30% w/w. The alkali-swollen material was poorly crystalline and was regenerated as alpha-chitin on washing with water. This conversion caused total collapse of the original microfibrillar morphology. These features are similar to those of 7 N-8 N HCl treatment reported earlier, but alkali treatment was free from depolymerization or deacetylation.     

Chemical composition and characterization of a galactomannoglucan from Gliocladium viride wall material

The following fractions were obtained from the wall material of Gliocladium viride : F1 (27.5%), a glucan, containing xylose, mannose and galactose, coluble in 1 M NaOH at 20°C; F2 (6.7%), a β-glucan-chitin complex, solubilized with 1 M NaOH at 20°C from the previous residue left overnight at −20°C; F3 (8.1%), a glucan, containing mannose and galactose solubilized with 1 M NaOH at 70°C; and F4, the insoluble residue, a β-glucan-chitin complex similar to F2, amounting to 31.3% of the wall material.
F1 was extracted with distilled water. The soluble material (F1S) was a galactomannoglucan (54.7%) and the inscluble (F1P) a glucan (45.3%). Periodate oxidation revealed the presence of glycerol, erythritol, threitol, ribitol, arabitol, mannose, galactose and glucose in F1S, and glycerol and glucose as the main components in F1P. The fractions obtained when F1S was purified through Sepharose CL6B, were methylated.