Cloning and Expression of an Endo-1,6-β-D-glucanase Gene (neg1) from Neurospora crassa

A gene (neg1) encoding an endo-1,6-β-D-glucanase from Neurospora crassa was cloned. The putative neg1 was 1443-bp long and encoded a mature endo-1,6-β-D-glucanase protein of 463 amino acids and signal peptide of 17 amino acids. The purified recombinant protein (Neg1) obtained from Escherichia coli showed 1,6-β-D-glucanase activity. No genes similar in sequence were found in yeasts and fungi.     

Toxins in Blast-diseased Rice Plants

The occurence of tenuazonic acid (T.A.), which had been isolated from the culture broth of blast fungus, in blast-diseased rice plants was surveyed to ascertain whether or not this substance is one of the vivotoxins. T.A. was detected in four of six samples of blast-diseased rice plants, two of which had relatively high T.A. contents; 379 and 91 mg per kg of the samples (dry weight).

Besides T.A., coumarin, o-coumaric acid and piricularin were also isolated from blast-diseased rice plants. The molecular formula of the last substance, which was tentatively presented in a previous paper, was corrected to C₁₈H₃₀N₂O₅ from the results of high resolution mass spectrometry.     

Numerical simulation and deacidification of nanomagnetic enzyme conjugate in a liquid–solid magnetic fluidized bed

The conventional deacidification method is difficult to achieve a better refining effect due to the high acid value in the rice bran crude oil, and the enzymatic esterification deacidification method can effectively reduce the acid value without generating chemical waste. In this study, the free lipase was immobilized on a magnetic polymer carrier Fe₃O₄/SiOx-g-P (GMA: glycidyl methacrylate) to obtain a immobilized lipase with a particle size of 105.30 ± 1.1 nm and an enzyme activity of 6580 ± 9.6 PLU/g (PLU: enzyme activity unit). Based on the batch deacidification process parameters, a multi-stage magnetic fluidized bed continuous circulation deacidification system was designed, and then the motion law of nanomagnetic immobilized lipase particles in liquid–solid magnetic fluidized bed was simulated by computer. When the iterative step was 5 × 10⁻⁵ s, the open porosity of the porous plate was 35.0%, the rice bran oil flow rate was 3.0 mm/s, and the magnetic field strength was 25.0 mT, which was beneficial to the deacidification reaction of rice bran oil. Under the conditions of magnetic immobilized lipase dosage of 4.0%, the phytosterol dosage of 22.0%, the molecular sieve dosage of 10%, the esterification temperature of 78.0 °C and the FFA (free fatty acid) content in rice bran oil decreased to 1.5%, after 48 h of reaction. The conversion rate is 92.8%, which provides a theoretical basis for the subsequent guidance of magnetic fluidized bed enzymatic continuous deacidification.     

Durum wheat by-products as natural sources of valuable nutrients

This review reports the use of wheat milling by-products for the extraction of high quality oil and vitamin E including our results on the exploitation of durum wheat bran as a valuable source of important healthful compounds. Wheat oil can be used as an ingredient in food, pharmaceutical or cosmetic preparations because it contains important bioactive compounds such as vitamin E, carotenoids and unsaturated fatty acids. Different methods are used for oil recovery from plant materials, such as solvent extraction, mechanical pressing or the eco-friendly supercritical carbon dioxide (SC-CO₂) extraction technology. By using SC-CO₂, we obtained an oil from durum wheat (Triticum durum Desf.) bran and optimized the extraction conditions to increase oil and vitamin E yields. Wheat bran, which is composed of pericarp, aleurone layer and germ, is discarded during the early stages of durum wheat milling processes to obtain a final product (semolina) that is stable over time. Maximum oil and vitamin E yields were obtained when a durum wheat bran matrix with particle size of ~30 mesh and a moisture content of 2.6 % was used. The optimal conditions for oil extraction were: 300–350 bar, 60–70 °C, and 4 l min^?1 gaseous CO₂ flow rate for 1 h. The chemical composition (vitamin E forms, carotenoids, quinones, lipids and fatty acids) of the SC-CO₂ extracted oil was analyzed and compared to that of the oil extracted by Soxhlet using hexane as solvent. The findings here reported highlight the importance of durum wheat bran as a rich source of valuable natural nutrients.     

Mechanism of rice bran lipase inhibition through fermentation activity of probiotic bacteria

Rice bran oil is known as wonder oil and it is the most important vegetable oil in Asia. Rice bran oil is extracted from bran that is the outer hard layer of rice. It is an emerging category in edible oil with a lot of nutritional properties and health benefits. Rice bran oil is heart-friendly, boosts up immunity, and prevents from other diseases occurring commonly in Pakistan. The current study aimed to stabilize rice bran oil through different probiotic isolates and to assess the nutritional content of rice bran oil after stabilization. The study was aimed to inactivate naturally occurring lipases that can hydrolyze oil into glycerol and free fatty acid which is a serious problem that gives it a rancid taste and smell. Antilipase activity was used to inactivate naturally occurring lipases that are a huge threat to the stabilization process. The fermentation process utilizes antilipase activity without affecting the nutritional value of oil. Lactobacillus strains were used for the stabilization of rice bran oil. Rice bran oil was extracted in the Soxhlet apparatus. The probiotic lab isolates Lactobacillus delbrueckii S2, Lactobacillus casei S5 and Lactobacillus plantarum S13 were applied to it to increase its shelf life and prevent oxidative rancidity. The extraction temperature of rice bran oil was maintained above 40 °C to inhibit lipase activity. Rice bran oil samples were stored at refrigeration temperature to arrest lipase activity. Probiotics maintained acidic pH to keep oil stabilization. Qualitative analysis was done to confirm rice bran oil stabilization. Determination of Free Fatty Acid (FFA) and saponification value confirmed that oxidative rancidity of rice bran oil was controlled by probiotics. FFA count was less than 10% and Saponification Value (SV) was 180. GC analysis was performed to analyze the FFA profile. Gas Chromatography results have shown 3 fatty acids. Statistical analysis has shown non-significant effect on different incubation temperatures of Lactobacillus isolates. Among the biological methods of stabilization, the use of probiotics is a novel concept and recommended for commercial application.     

Biochemical Studies on Ricin Part X Effect of the Two Constituent Polypeptide Chains of Ricin D toward Mouse Sarcoma Ascite Tumor Cells

A maltotetraose-forming amylase from Pseudomonas stutzeri was highly purified by adsorption on starch granules and by chromatographies on Sephadex G-100 and DEAE-cellulose. The purified enzyme showed a single band in polyacrylamide gel electrophoreses with or without sodium dodecylsulfate. The optimum pH for enzyme action on starch was 6.0-6.5, and the optimum temperature was 45°C. The purified enzyme attacked starch from the non-reducing end to produce α-anomer oligosaccharides. This indicated that the enzyme was an exo-α-amylase which had not hitherto been found. The enzyme activity was markedly inhibited by the addition of Cu²⁺, Hg²⁺, N-bromosuccinimide and 2,3-butanedione. The molecular weight of the enzyme determined by the method of Weber and Osborn was about 5.7 × 10⁴. The isoelectric point of the enzyme was estimated to be 5.3 by polyacrylamide gel electrofocusing. The Km and k₀ values of this enzyme for starch, glycogen, short chain amylose and some maltooligosaccharides were calculated from Lineweaver-Burk plots.     

Biodiesel production from rice bran by a two-step in-situ process

The production of fatty acid methyl esters (FAMEs) by a two-step in-situ transesterification from two kinds of rice bran was investigated in this study. The method included an in-situ acid-catalyzed esterification followed by an in-situ base-catalyzed transesterification. Free fatty acids (FFAs) level was reduced to less than 1% for both rice bran A (initial FFAs content = 3%) and rice bran B (initial FFAs content = 30%) in the first step under the following conditions: 10 g rice bran, methanol to rice bran ratio 15 mL/g, H₂SO₄ to rice bran mass ratio 0.18, 60 °C reaction temperature, 600 rpm stirring rate, 15 min reaction time. The organic phase of the first step product was collected and subjected to a second step reaction by adding 8 mL of 5 N NaOH solution and allowing to react for 60 and 30 min for rice bran A and rice bran B, respectively. FAMEs yields of 96.8% and 97.4% were obtained for rice bran A and rice bran B, respectively, after this two-step in-situ reaction.     

Biodiesel production from rice bran oil and supercritical methanol

In this study, production of biodiesel from low cost raw materials, such as rice bran and dewaxed-degummed rice bran oil (DDRBO), under supercritical condition was carried out. Carbon dioxide (CO₂) was employed as co-solvent to decrease the supercritical temperature and pressure of methanol. The effects of different raw materials on the yield of biodiesel production were investigated. In situ transesterification of rice bran with supercritical methanol at 30 MPa and 300 °C for 5 min was not a promising way to produce biodiesel because the purity and yield of fatty acid methyl esters (FAMEs) obtained were 52.52% and 51.28%, respectively. When DDRBO was reacted, the purity and yield were 89.25% and 94.84%, respectively. Trans-FAMEs, which constituted about 16% of biodiesel, were found. They were identified as methyl elaidate [trans-9], methyl linoleaidate [trans-9, trans-12], methyl linoleaidate [cis-9, trans-12], and methyl linoleaidate [trans-9, cis-12]. Hydrocarbons, which constituted about 3% of the reaction product, were also detected.     

Optimization of supercritical carbon dioxide extraction of silkworm pupal oil applying the response surface methodology

Supercritical carbon dioxide extraction (SC-CO₂) of oil from desilked silkworm pupae was performed. Response surface methodology (RSM) was applied to optimize the parameters of SC-CO₂ extraction. The effects of independent variables, including pressure, temperature, CO₂ flow rate, and extraction time, on the yield of oil were investigated. The statistical analysis showed that the pressure, extraction time, and the quadratics of pressure, extraction time, and CO₂ flow rate, as well as the interactions between pressure and temperature, and temperature and flow rate, showed significant effects on oil yield. The optimal extraction condition for oil yield within the experimental range of the variables researched was at 324.5 bar, 39.6 °C, 131.2 min, and 19.3 L/h. At this condition, the yield of oil was predicted to be 29.73%. The obtained silkworm pupal oil contained more than 68% total unsaturated fatty acids, and alpha-linolenic acid (ALA) accounted for 27.99% in the total oil.     

Rice bran stabilization and rice bran oil extraction using ohmic heating

Ohmic heating has been shown to increase the extraction yields of sucrose from sugar beets, apple juice from apples, beet dye from beet root, and soymilk from soybeans. Rice bran is a byproduct of the rice milling process that has economic potential by virtue of highly nutritious rice bran oil contained within the bran. In this study, ohmic heating was used to stabilize rice bran and to improve rice bran oil extraction yield as compared to microwave heating and a control (no heating). Results showed that ohmic heating is an effective method for rice bran stabilization with moisture addition. Free fatty acid concentration increased more slowly than the control for raw bran samples subjected to ohmic heating with no corresponding temperature rise, indicating that electricity has a non-thermal effect on lipase activity. Ohmic heating increased the total percent of lipids extracted from rice bran to a maximum of 92%, while 53% of total lipids were extracted from the control samples. Lowering the frequency of alternating current significantly increased the amount of oil extracted, probably due to electroporation. Ohmic heating was successfully applied to rice bran despite its high oil content. This could have important implications for the enhanced extraction of non-polar constituents.     

Cellulase Production in Semisolid Cultures of Trichoderma viride

Cellulase was produced by Trichoderma viride in semisolid cultures of rice bran, rice straw and rice hulls. T. viride QM 9414 generally produced higher cellulolytic activity on CM-cellulose (C_x activity) using rice bran-rice hull mixture (2:1 w/w) as substrate compared to strains ITCC 1433 and D 4014. It showed higher C_x activity on rice bran-rice straw mixtures than on rice bran-rice hull mixtures. Maximal extraction of the enzyme from mold bran was obtained with 0.05 m sodium citrate buffer, pH 3.5.     

Biochemical characterization of a 27 kDa 1,3-β-d-glucanase from Trichoderma asperellum induced by cell wall of Rhizoctonia solani

Trichoderma asperellum produces two extracellular 1,3-β-d-glucanase upon induction with cell walls from Rhizoctonia solani. A minor 1,3-β-d-glucanase was purified to homogeneity by ion exchange chromatography on Q-Sepharose and gel filtration on Sephacryl S-100. A typical procedure provided 13.8-fold purification with 70% yield. SDS-PAGE of the purified enzyme showed a single protein band of molecular weight 27 kDa. The enzyme exhibited optimum catalytic activity at pH 3.6 and 45 °C. It was thermostable at 40 °C, and retained 75% activity after 60 min at 45 °C. The K_m and V_max values for 1,3-β-d-glucanase, using laminarin as substrate, were 0.323 mg ml⁻¹ and 0.315 U min⁻¹, respectively. The enzyme was strongly inhibited by Hg²⁺ and SDS. The enzyme was only active toward glucans containing β-1,3-linkages. Peptide sequences showed similarity with two endo-1,3(4)-β-d-glucanases from Aspergillus fumigatus Af293when compared against GenBank non-redundant database.     

An Asymmetric Synthesis of 2-Substituted Piperidines through Ozonolysis of Cyclopentenes and Reductive Aminocyclization

By the action of ozone, sodium cyanoborohydride and the optically active benzylic amines 2, the 1-substituted cyclopentenes 1, 5 and 9 were converted to a diastereoisomeric mixture of 1,2-disubstituted piperidines (3, 6 and 10), respectively. Hydrogenation of these compounds and the following work-up yielded optically active 2-alkylpiperidines (4, up to 68% e.e.), pipecolic acid (7, 84%e.e.) and 2-(hydroxymethyl)piperidine (11, up to 85%e.e.). Chromatographic separation of the major isomers of 3b and 6 enabled optically pure coniine (4b) and pipecolic acid (7) to be prepared, respectively.     

Production of mosquitocidal <Emphasis Type="Italic">Bacillus sphaericus</Emphasis> by solid state fermentation using agricultural wastes

In this study, Bacillus sphaericus NRC 69 was grown in culture media, in which 12 agricultural wastes were tested as the main carbon, nitrogen and energy sources under solid state fermentation. Of the 12 tested agricultural by-products, wheat bran was the most efficient substrate for the production of B. sphaericus mosquitocidal toxins against larvae of Culex pipiens (LC₅₀ 1.2 ppm). Mixtures of tested agricultural wastes separately with wheat bran enhanced the produced toxicity several folds and decreased LC₅₀ between 3.7- and 50-fold in comparison with that of agricultural wastes without mixing. The toxicity of B. sphaericus grown in wheat bran/rice hull at 8/2 (g/g) and wheat bran/barley straw at 1/4 (g/g) showed the same toxicity as that in wheat bran medium (LC₅₀ decreased 17- and 16-fold, in comparison with that in rice hull or barely straw media, respectively). In wheat bran medium, the maximum toxicity of the tested organism obtained at 50% moisture content, inoculum size 84 × 10⁶ CFU/g wheat bran and incubation for 6 days at 30°C. Addition of cheese whey permeate at 10% to wheat bran medium enhanced the toxicity of B. sphaericus NRC 69 about 46%.     

Stereoselective Synthesis of (S,E)-6-Isopropyl-3,9-dimethyl-5,8-decadienyl Acetate,the (S)-Enantiomer of the Yellow Scale Pheromone

The (S)-enantiomer of the sex pheromone of the yellow scale (Aonidiella citrina), (S,E)-6-isopropyl-3,9-dimethyl-5,8-decadienyl acetate, was stereoselectively synthesized from (R)-(+)-citronellic acid.     

Trypsin inhibitor,mineral availability,and performance of broiler chickens fed on diets based on rice bran

Trypsin inhibitor in rice bran was inactivated by moist heat, but not by dry heating or treatments with 0.1% H₂SO₄, 0.1% NaOH or 0.1% NaCl. Heating rice bran could not improve its growth promotion value in the diet of broilers, which was related to the feed intake. Groundnut meal was the major protein supplement in the diets. Rice bran diets did not affect weight of pancreas (1.45–1.50 g/kg body weight). Use of ⁴⁵Ca revealed that availability of calcium on a rice bran diet was lower than on a maize-based diet. A similar effect on ⁵⁹Fe-availability was noted, although this result needs confirmation. Lack of differences in weight of thyroid glands and uptake of ¹³¹I by thyroids of birds on maize- and rice bran-based diets indicated the absence of any goitrogenic substance in the rice bran.     

Extraction of Oils from Wheat Germ with Supercritical Carbon Dioxide

The separation of oil from wheat germ by extracting with supercritical carbon dioxide (C0₂) is described. The solubility of wheat germ oil in supercritical C0₂ at 200 atm and 40°C was about 0.35 weight%. The effect of pressure on the extraction process with liquid or supercritical C0₂ was of great significance. On the other hand, the effect of temperature on the extraction process was small. Oil extracted with supercritical C0₂ was lighter in color and contained less phosphorus than that extracted with hexane. The contents of α- and β-tocopherol in the oil extracted with supercritical CO₂. were comparable to those in the hexane-extracted oil     

Components of Unsaponifiable Matter of Rice Bran Oil

Components of the unsaponifiable matter of crude rice bran oil were examined. A new compound having melting point of 157~158°C was obtained. Molecular weight was determined and ultraviolet and infrared analyses were carried out. Squalene and ferulic acid ester were confirmed to exist in the unsaponifiablc matter of crude rice bran oil. Quantitative relation of components was calculated and found to be 42% sterols, 24% higher alcohols. 20% ferulic acid esters, 10% hydrocarbons and 2% unreported compound.     

Isolation of a Thiamine-binding Protein from Rice Germ and Distribution of Similar Proteins

A thiamine-binding protein was purified from rice germ (Oryza sativa L.) by extraction, salting-out with ammonium sulfate, and column chromatography. From the results of molecular mass, K_d and B_max values for thiamine-binding, binding specificity for thiamine phosphates and analog, the protein was suggested to be identical to the thiamine-binding protein in rice bran. The thiamine-binding protein w as more efficiently purified from rice germ than from rice bran. The protein was rich in glutamic acid (and/or glutamine) and glycine. The protein did not show immunological similarity to thiamine-binding proteins in buckwheat and sesame seeds. However proteins similar to the thiamine-binding protein from rice germ existed in gramineous seeds. They were suggested to have thiamine-binding activity and to be of the same molecular mass as the thiamine-binding protein.     

Process optimization for extraction of carotenoids from shrimp waste with vegetable oils

Shrimp waste is an important source of natural carotenoid. Studies were carried out to determine the extraction yield of shrimp waste carotenoids in different vegetable oils. Highest yield was obtained by extraction using refined sunflower oil compared to groundnut oil, gingelly oil, mustard oil, soy oil, coconut oil and rice bran oil. The extraction yield of carotenoids in sunflower oil was significantly influenced by level of oil to waste (p < 0.05), time (p < 0.01) and temperature (p < 0.001) of heating waste with oil before centrifugation to separate pigmented oil. A regression equation was derived for carotenoid yield as a function of time of heating, temperature of heating and oil level to waste. The optimized conditions for extraction of shrimp waste carotenoids in sunflower oil were determined to be oil level to waste of 2, temperature of 70 degrees C and heating time of 150 min.