Body Condition Score, Rumination, Intake, Milk Production and Milk Composition of Grazing Dairy Cows Supplemented with Rumen-Protected Lysine and Methionine

The study utilised a pasture grazing based, voluntary traffic automatic milking system to investigate milk production of cows fed a pasture-based diet and supplemented with a pellet formulated with vs. without rumen-protected lysine and methionine (RPLM). The study adopted a switch-over design (over two periods of 5 and 10 weeks, respectively) and used 36 cows and equally allocated them into two experimental groups. The RPLM (Trial) pellet had 2% lower crude protein, but similar metabolizable energy content compared to the Control pellet. Pellet intake was 10.0 and 9.4 kg/day/cow. Milk yield was 36.2 and 34.4 kg/day/cow (p = 0.23), and energy corrected milk was 35.1 and 33.8 kg/day/cow (p = 0.076), and milk solids was 2.55 and 2.46 kg/cow/day (p = 0.073) in the Control and Trial groups, respectively. Milk fat%, milk protein%, milk fat: protein ratio, milking frequency and rumination time were not different between the two groups (p > 0.05). In period 1, plasma glucose was 3.1 mmol/L for both groups and milk urea were 150 and 127 mg/L in the Control and Trial groups, respectively. Both plasma glucose (as a proxy for energy supply) and milk urea (as a proxy for nitrogen use efficiency; NUE) were not different between groups (p > 0.05). This study showed that under a grazing pasture system, feeding lactating dairy cows a low protein pellet with RPLM supplementation, maintained milk production performance and NUE, compared with cows fed a high protein Control pellet diet with no RPLM. Further research should assess the long-term (seasonal) effects of feeding a diet formulated with RPLM on cow intake, health and reproductive performance.

Proteomic and Metabolomic Analyses of Maggots in Porcine Corpses for Post-Mortem Interval Estimation

Post-mortem interval (PMI) estimation is a critical task in forensic science. In this study, we used maggots collected from pig carcasses and applied an integrated proteomics and metabolomics approach to determine potential candidate substances for the estimation of PMI. After methanol precipitation, the supernatant containing metabolites and the protein pellet were separated and subjected to metabolomic and proteomic analyses using liquid chromatography-tandem mass spectrometry (LC-MS/MS). MS/MS data were analyzed for identification and quantification using Proteome Discoverer and Compound Discoverer software. A total of 573 metabolites and more than 800 porcine proteins were identified in maggots. This is the first dataset of proteins and metabolites in maggots collected from porcine carcasses. In this study, guanosine monophosphate, xanthine, inosine, adenosine, and guanine were detected with a similar tendency to increase during early days of maggot development and then decreased gradually. We broadly profiled various biomolecules through analysis in the spot of incident. Especially, we confirmed that proteome and metabolome profiling could be performed directly and indirectly.

51 Observing the Effect of Creep Feeding on Calf Behavior Before and After Weaning

The objective was to determine the effect of creep feeding on calf behavior before and after weaning. Cow-calf pairs (n = 54) were stratified by cow age, cow and calf BW, and allotted to 9 pastures. Pastures were randomly assigned to 3 treatments: unsupplemented control (CON), lower protein pellet (14.3% CP; LP), and a higher protein pellet (18.6% CP; HP) for 82 d. Pre-weaning calf behaviors were observed for 9 h on d 60, 64, and 74: position (standing or lying), location (pen, pasture, shade, creep), and consumption (creep, grass, milk, water, none). Post-weaning calf behaviors were observed for 12 h on d 83 and 84: position, walking, drinking, eating, and vocalizations. Prior to weaning, an interaction (P £ 0.01) occurred for consumption behaviors; CON calves spent the greatest percentage of time grazing at h 7 and 8 and not consuming at h 11. Also, LP calves had the greatest percentage of time nursing at h 8 and 11, with CON calves being intermediate, and HP calves being the least. On the day after weaning (d 83), an interaction (P = 0.02) was observed for time spent eating; CON calves spent the greatest time eating at h 10 and LP calves were the greatest at h 11. An interaction (P < 0.01) was observed for time spent walking on d 83; HP calves spent the greatest percentage of time walking at h 9 and 13. On d 84, an interaction (P < 0.01) was observed for time eating on d 84; CON calves spent the greatest percentage of time eating at h 7, 9, 11, 14, and 16 as well as overall (P = 0.05). An interaction (P < 0.01) was observed for number of vocalizations on d 84; CON calves had the greatest number of vocalizations at h 16–18 as well as overall (P < 0.01). Overall, creep feeding altered calf behaviors before and after weaning.

High-yield skeletal muscle protein recovery from TRIzol after RNA and DNA extraction

Extraction of DNA, RNA and protein from the same sample would allow for direct comparison of genomic, transcriptomic and proteomic information. Commercially available kits exhibit poor protein yield and the TRIzol® reagent produces a protein pellet that is extremely difficult to solubilize. In response to these limitations, this study presents an optimized method for the extraction of protein from the organic phase of TRIzol that allows for higher yield recovery of skeletal muscle protein compared with direct homogenization in a common protein lysis buffer. The presented method is inexpensive, simple and fast, requires no additional treatment of the protein pellet for dissolution, and is compatible with downstream western blot applications.

Optimization of a Colorimetric Assay to Determine Lactate Dehydrogenase B Activity Using Design of Experiments

Lactate dehydrogenase B (LDH-B) is overexpressed in lung and breast cancer, and it has been considered as a potential target to treat these types of cancer. Herein, we propose a straightforward incomplete factorial (IF) design composed of 12 combinations of two reaction buffers, three pH values, three salt (NaCl) concentrations, and three incubation times, which we called IF-BPST (Buffer/pH/Salt/Time), for the optimization of a colorimetric LDH-B assay in a final volume of 100 µL using 96-well plates. The assay is based on the absorbance change at ~570 nm and the color change of the reaction mixture due to the release of NADH that reacts with nitroblue tetrazolium (NBT) and phenazine methosulfate (PMS), resulting in the formation of a blue-purple formazan. The results obtained using the IF-BPST were comparable with those obtained by response surface methodology. Our work revealed that the NBT/PMS assay with some modifications can be used to measure the activity of LDH-B and other dehydrogenases in a high-throughput screening format at the early stages of drug discovery. LDH-B containing lysates cannot be assayed directly, however, due to the sensitivity of the method toward detergents. Thus, we suggest precipitating the proteins in the lysates to remove the interfering detergents, and then to dissolve the protein pellet in a suitable buffer and carry out the assay.

Improving the pre-weaning nutrition of calves by supplementation of the cow and/or the calf while grazing low quality pastures. 1. Cow production

Large numbers of weaner calves destined for meat production in Australia are born to cows grazing low quality pastures and have low growth rates to weaning. A study was undertaken in a New South Wales subtropical coastal area (Grafton, 29°38′S, 152°54′E) to increase these growth rates by supplementing cows to mid lactation and/or their calves up to weaning. This paper reports on the effects of these supplementation regimes on cow production. Cows were either not supplemented or supplemented with cottonseed meal twice-a-week at 4.2 kg/head over 145 days, commencing before the expected start of calving. Calves, depending on their treatment group, either had unrestricted access to a ‘creep’ containing a high-energy, high-protein pellet given from 75 or 150 days old, or had no access at all to ‘creep’ feed. However, all cows and their calves had access to mineral blocks. Cottonseed meal-supplemented cows were heavier (408 v. 362 kg) and had higher condition scores (3.3 v. 2.9 units) than non-supplemented cows when weighed and scanned in November 1996, after all cows had calved, and they maintained these differences until April 1997 when calves were weaned. There was a trend for cows, whose calves had access to a ‘creep’ from 150 days old, to gain weight towards the end of lactation whereas cows from other treatment groups lost weight. Non-supplemented cows had a 3-fold greater daily intake of mineral blocks than cottonseed meal cows (178 v. 61 g/cow) providing them with an additional 9.8 g phosphorus/cow. Cottonseed meal cows were observed to have fewer grazing incidences (60 v. 75% of activities) during supplementation than non-supplemented cows, with grazing instances increasing for all cows during lactation. Cottonseed meal cows whose calves had access to ‘creep’ feed continued to have fewer grazing incidences (74 v. 90%) than other cows 70 days post-supplementation. Milk yield was higher in cottonseed meal cows than in non-supplemented cows (4.8 v. 4.0 kg/day). Following a synchronised insemination program, at the end of cottonseed meal supplementation, the pregnancy rate of formerly cottonseed meal-supplemented cows was twice that (40 v. 20%) of non-supplemented cows. The combination of cottonseed meal supplements and macrominerals from blocks improved cow liveweight, condition score, milk yield and fertility, and was a successful strategy for increasing production from low quality pastures.

A veterinary model for the induction of primary nutritional ketosis in dairy cattle

A method for inducing primary nutritional ketosis in dairy cattle was developed in order to evaluate a novel technique for the monitoring of the development of ketosis through to clinical disease state. Six freshly calved, multiparous Holstein-Friesian cows were given a 500 g/kg crude protein pellet in small but frequent meals through an out-of-parlour feeder in conjunction with an allowance of grass silage restricted to 20 kg wet weight per day. Development of the disease was monitored through clinical signs, body condition scoring, weight change measures and metabolic profiling of plasma samples with particular reference to beta-hydroxybutyrate, non-esterified fatty acid and glucose levels. Four animals within the disease model group became clinically ketotic with the remaining two animals showing signs of subclinical ketosis. A further four newly calved cows, which were maintained on a normal production ration to act as a control group, did not develop signs of ketosis.

The effect of feeding varying levels of barley and dried molassed sugar beet feed compared to a commercial creep on the performance of intensively reared Iambs

Feed is the major cost of indoor lamb production, it accounts for up to £12/lamb reared (MLC, 1994) which is equivalent to 23% of the output value. Most intensively produced lambs are fed expensive specially formulated lamb creep diets. The aim of this experiment was to evaluate the effect varying the proportions of the starch (barley, WB) and digestible fibre (dried molassed sugar beet feed; SBF) and a protein pellet compared to a commercial lamb creep (LC) on lamb physical and financial performance.180 early weaned lambs, (weaned at 6 weeks of age), weighing on average 17.5 kg liveweight were allocated to one of six treatments; 0% SBF, 20% SBF, 40% SBF, 60% SBF, 80% SBF and LC. Each diet contained 20% protein balancer pellets with the remainder being whole barley. The LC diet was offered alone.

The effect of feeding varying levels of barley and dried molassed sugar beet feed compared to a commercial creep on the performance of intensively reared Iambs

Feed is the major cost of indoor lamb production, it accounts for up to £12/lamb reared (MLC, 1994) which is equivalent to 23% of the output value. Most intensively produced lambs are fed expensive specially formulated lamb creep diets. The aim of this experiment was to evaluate the effect varying the proportions of the starch (barley, WB) and digestible fibre (dried molassed sugar beet feed; SBF) and a protein pellet compared to a commercial lamb creep (LC) on lamb physical and financial performance.180 early weaned lambs, (weaned at 6 weeks of age), weighing on average 17.5 kg liveweight were allocated to one of six treatments; 0% SBF, 20% SBF, 40% SBF, 60% SBF, 80% SBF and LC. Each diet contained 20% protein balancer pellets with the remainder being whole barley. The LC diet was offered alone.

COVALENT BINDING OF CYCLOOXYGENASE AND LIPOXYGENASE PRODUCTS TO HUMAN PLATELET PROTEINS

Several studies in acellular systems have shown a covalent binding of eicosanoids to proteins (1). We have therefore investigated whether eicosanoids bind covalently to proteins in intact platelets. After incubation of washed human platelets with 14C-arachidonic acid, ethanol precipitation followed by extractions, a small fraction of the radioactivity (0.3%) was tightly bound to the protein pellet. Four criteria suggest the covalent nature of this binding. The radioactivity remained bound after exhaustive extractions with solvents of various polarities, and was not removed by dialysis against SDS-buffer. 15 labelled protein bands could be separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Finally, exhaustive enzymatic hydrolysis of platelet proteins by several proteases liberated an amphipathic radioactive material which had a chromatographic behaviour similar to that of a known peptidolipid, leuko-triene c4 This covalent binding involved products of both cyclooxygenase and lipoxygenase pathways : it was partially inhibited by indomethacin (±40%) and completely abolished by eicosatetraynoic acid. The covalent binding was increased five-fold by dazoxiben, suggesting attachment of prostaglandin endoperoxydes , and by diamide (a glutathione depleting agent) (2) suggesting the involvement of 12-hydro-peroxyeicosatetraenoic acid. Dazoxiben and diamide intensified selectively the labelling of distinct protein bands, separated by SDS-PAGE. Further studies will be needed to evaluate the possible physiological significance of this covalent modification.(1) Wilson, A.G.E. et al. Prostaglandins 18: 409-422, 1979.(2) Bryant, R.W. et al., J. Biol. Chem. 257: 14937-14943, 1982