Towards non-invasive methods to determine the effect of treatment of soya-bean meal on lysine availability in dairy cows

Lysine (Lys) availability in three different soya-bean meal (SBM) products was determined using the following techniques: whole body (WB) net flux of Lys, digestible Lys (duodenal flow × intestinal digestibility) and the plasma Lys response curve method of Rulquin and Kowalczyk (2003). Four multiparous Holstein cows (173 days in milk) were equipped with ruminal and duodenal cannulas and used in a 4 × 4 Latin square experiment with 14-day periods. The animals were fed either solvent-extracted SBM (SE), expeller-processed SBM (EP) or lignosulphonate-treated SBM (LS) at 23% of the diet dry matter (DM). The fourth treatment (SE70) consisted of a continuous infusion of Lys (70 g/day) into the omasum of cows fed the SE diet. Chromium(III) oxide was included as a digesta marker in order to determine the duodenal flow of amino acids (AA). On day 12 of each experimental period, six blood samples were collected to determine plasma Lys concentrations. Immediately after that, a pulse dose of L-[2-15N] Lys was administered in the jugular vein. Jugular blood samples were then collected at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 13, 16, 19, 25 and 31 min after the injection to determine 15N Lys enrichment. On each of days 13 and 14, eight digesta samples were collected and pooled by period. Amongst the diets of SBM (SE, EP, LS), no differences were observed for duodenal Lys flow or digestible Lys. Duodenal flow of microbial N with SE was numerically higher, compared with EP and LS, indicating enhanced duodenal supply of microbial Lys for this diet, and this may have compensated for the additional Lys derived from undegradable protein in rumen-protected SBM products (EP and LS). The use of the plasma response curve method as well as the measurement of WB Lys flux also revealed no differences in Lys availability among the SBM products. The WB flux method resulted in 100% post-ruminal recovery of the Lys infused with diet SE70 compared with the control diet SE, which indicates that the method is reliable for determining Lys availability. The Lys flux approach not only allows for the estimation of intestinally available essential AA but also it avoids the use of cannulated animals.     

Caecal fermentation characteristics, blood composition and growth of rabbits on substitution of soya-bean meal by unconventional high-glucosinolate mustard ( Brassica juncea) meal as protein supplement

Effect of graded levels of high-glucosinolate mustard (Brassica juncea) meal as substitute of soya-bean meal (SBM) in broiler rabbit diets was studied. Forty weaning rabbits of Soviet Chinchilla and White Giant breed were randomly allocated to one of four experimental diets containing mustard meal (MM) 0, 80, 160 and 245 g/kg. The experiment lasted for 8 weeks. MM had 54.8 mg total glucosinolates (TGLSs) per g dry matter (DM). Diets had TGLS 3.8, 8.4 and 11.98 mg/g DM in 80, 160 and 245 g MM diets, respectively. MM-incorporated diets had higher digestible and linearly (P < 0.01) higher metabolisable energy (ME) content. However, the effect on total tract apparent digestibility of DM, and crude protein was quadratic. Average daily gain (ADG) reduced (P < 0.05) linearly with increasing MM levels in diet, still 80 and 160 g MM diets had similar ADG compared to that of SBM diet. Caecum weight reduced linearly (P < 0.05) with increasing MM levels in diet. The pH of caecal content ranged between 5.85 and 6.19, total N between 1.19 and 1.48 (g per 100 g) and total volatile fatty acids between 4.7 and 5.8 mmol per 100 g, and they were not statistically different. NH3-N ranged between 31.2 and 39.0 mg per 100 ml, and reduced linearly (P < 0.05) while trichloroacetic acid-precipitable nitrogen increased linearly (P < 0.01, ranged between 114 and 247 mg per 100 ml) with increasing MM levels in diet. Blood haemoglobin, packed cell volume and lymphocytes were higher (quadratic effects, P < 0.05) on 245 MM diet, whereas white blood cell count reduced linearly (P < 0.01). Serum aspartate aminotransferase increased linearly (P < 0.01) while alanine aminotransferase and alkaline phosphatase activity, protein, erythrocytes sedimentation rate and red blood cell counts were not affected by MM. Serum Cu, Na and K content increased linearly (P < 0.05) with increasing MM levels. Liver Cu concentration showed quadratic (P < 0.05) increase. Rabbits tolerated 8.4 mg TGLS per g diet (160 g MM per kg) during active growth without any apparent effect on health and growth. It is concluded that MM can replace up to 66% SBM protein in rabbit feeding, whereas complete replacement of SBM with MM reduced feed intake and ADG by 23% and 13%, respectively. Further studies are required to confirm these inclusion levels and glucosinolate tolerance of rabbits.     

Response surface optimization of fermentation conditions for producing xylanase by <Emphasis Type="Italic">Aspergillus</Emphasis><Emphasis Type="Italic">niger</Emphasis> SL-05

Fermentation conditions were statistically optimized for producing extracellular xylanase by Aspergillus niger SL-05 using apple pomace and cotton seed meal. The primary study shows that culture medium with a 1:1 ratio of apple pomace and cotton seed meal (carbon and nitrogen sources) yielded maximal xylanase activity. Three significant factors influencing xylanase production were identified as urea, KH(2)PO(4), and initial moisture content using Plackett-Burman design study. The effects of these three factors were further investigated using a design of rotation-regression-orthogonal combination. The optimized conditions by response surface analysis were 2.5% Urea, 0.09% KH(2)PO(4), and 62% initial moisture content. The analysis of variance indicated that the established model was significant (P < 0.05), "while" or "and" the lack of fit was not significant. Under the optimized conditions, the model predicted 4,998 IU/g dry content, whereas validation experiments produced an enzymatic activity of xylanase at 5,662 IU/g dry content after 60 h fermentation. This study innovatively developed a fermentation medium and process to utilize inexpensive agro-industrial wastes to produce a high yield of xylanase.     

Selection and characterization of feather-degrading bacteria from canola meal compost

Canola meal that contains a high level of protein (40% crude protein) was used as compost material for the isolation of feather-degrading bacteria. After 7 and 14 days, bacteria were isolated from compost amended and unamended with soil. Eighty bacterial isolates from canola meal compost were then grown on milk-agar and isolates that produced proteolytic enzymes were identified by the formation of clear haloes around the colonies. A feather medium was chosen for a secondary selection of feather-degrading isolates. Of the eight isolates that hydrolyzed milk protein, five isolates hydrolyzed feathers. Their keratinolytic activities were subsequently confirmed by an assay using azo-keratin as substrate. Seven of the eight bacteria that hydrolyzed milk protein were Bacillus spp, and all five isolates that hydrolyzed feathers were strains of Bacillus licheniformis. Protease inhibition studies indicated that serine proteases are the predominant proteolytic enzymes produced by these feather-degrading isolates. Received 02 April 1999/ Accepted in revised form 17 June 1999     

Promotion of plant and root growth by soybean meal degradation products

The growth of Brassica campestris, Solanum tuberosum L., Lycopersicum esculentum and Brassica junce in the field were promoted by the degraded soybean meal products (DSP). The root hair number of Brassica campestris was increased when 10 l DSP (containing 30 mg peptides + amino acids ml^–1) were added to 10 ml plant growth medium. A chemical fertilizer and an acid-hydrolyzed DSP did not show such an effect.     

Biodegradation of glucosinolates in brown mustard seed meal (Brassica juncea) by Aspergillus sp. NR-4201 in liquid and solid-state cultures

Aspergillus sp. NR-4201 was assessed by degrading glucosinolates in brownmustard seed meal (Brassica juncea). A liquid culture of the strain, in a medium derived from the meal, produced total degradation of glucosinolates at 32 h. Under these conditions, the glucosinolate-breakdown product, allylcyanide, was formed inculture filtrates. In a plate culture under sterile conditions, the growth of the strain inheat-treated meal media was shown to be effective at 30 °C with 51% moisture,as determined by the measurement of the colony growth rate. On the laboratory scale,solid-state culture under the same conditions gave rise to total glucosinolate degradationwithin 48 h. In comparison, under non-sterile conditions in either heat-treated or nonheat-treated meal samples, the degradations were complete after 60 and 96 h, respectively.In these cases, growth was associated with some out-growths of contaminating fungi,mainly Rhizopus sp. and Mucor sp. The glucosinolate-breakdown product,allylcyanide, was not detected in the solid-state meal-media culture presumably due toevaporative loss from the fermentation matrix.     

Efficient transformation of the yellow fever mosquito Aedes aegypti using the piggyBac transposable element vector pBac[3xP3-EGFP afm

We report efficient germ-line transformation in the yellow fever mosquito Aedes aegypti accomplished using the piggyBac transposable element vector pBac[3xP3-EGFP afm]. Two transgenic lines were established and characterized; each contained the Vg-Defensin A transgene with strong eye-specific expression of the enhanced green fluorescent protein (EGFP) marker gene regulated by the artificial 3xP3 promoter. Southern blot hybridization and inverse PCR analyses of genomic DNA demonstrated a precise piggyBac-mediated, single copy insertion of the pBac[3xP3-EGFP afm,Vg-DefA] transposon in each transgenic line. For each line, genetic analysis confirmed stability and integrity of the entire transposon construct in the mosquito genome through the G₂–G₆ generations. Successful establishment of homozygous transgenic lines indicated that in both cases a non-lethal integration of the transposon into the mosquito genome had occurred. The 3xP3-EGFP marker was tested in mosquitoes with different genetic backgrounds. In white-eyed transgenic mosquitoes, the strong eye-specific expression of GFP was observed throughout all stages of development, starting from newly hatched first instar larvae to adults. A similar level and pattern of fluorescence was observed in red-eyed mosquitoes that were generated by crossing the 3xP3-EGFP transformants with the kh^w white-eye mosquitoes transformed with the Drosophila cinnabar gene. Importantly, the utility of the 3xP3-EGFP, as marker gene for transformation of wild type mosquitoes, was demonstrated by strong eye-specific GFP expression in larval and pupal stages of black-eyed hybrids of the 3xP3-EGFP white-eye transformants and the wild type Rockefeller/UGAL strain. Finally, analysis of the Vg-DefA transgene expression in transformants from two established lines demonstrated strong blood-meal activation and fat-body-specific expression regulated by the Vg 1.8-kb 5′ upstream region.     

Isolation and characterization of a small molecular weight protein of linseed meal

A small M_r, protein from linseed meal has been isolated by CM-Sephadex chromatography. The protein was found to be homogeneous by the techniques of gel filtration, polyacrylamide gel electrophoresis and ultracentrifugation. It had S_20,w value of 1.6S. Amino acid composition of the protein revealed a high amount of glutamic acid, cystine, arginine and glycine. The absorption spectrum of the protein consisted of a peak at 280 nm with a shoulder at 290 nm. The fluorescence emission maximum was at 340 nm. The protein contained large amounts of α-helix and β-structure. SDS-PAGE showed the protein to consist of a single polypeptide chain. The M_r estimated by Archibald's method, sedimentation-diffusion method and gel filtration was 17 000,16 000 and 15 000 respectively. Difference spectra studies as a function of pH and temperature showed no variation in the conformation of the protein, probably due to disulphide bridges.     

Early Time-Restricted Feeding Improves Insulin Sensitivity,Blood Pressure,and Oxidative Stress Even without Weight Loss in Men with Prediabetes

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