Abstract: An immunoglobulin G (IgG) was spray-dried on a Büchi 190 laboratory spray-dryer at inlet and outlet air temperatures of 130 and 190°C, respectively. The IgG solution contains initially 115mg/ml IgG plus 50mg/ml sorbitol. After dialysis, at least 80% of low molecular weight component was removed. After spray-drying the dialyzed IgG and immediate redissolution of the powder, an increase in aggregates from 1 to 17% occurred. A major shift towards increase β-sheet structure was detected in the spray-dried solid, which, however, reverted to native structure on redissolution of the powder. A correlation between aggregation determined by size exclusion chromatography and alterations in secondary structure determined by Fourier transformation infra-red spectroscopy could not therefore be established. On spray-drying a non-dialyzed, sorbitol-containing IgG only some 0.7% aggregates were formed. The sorbitol is therefore evidently able to stabilize partially the IgG during the process of spray-drying. Addition of trehalose to the liquid feed produced quantitatively the same stabilizing action on the IgG during spray-drying as did the sorbitol. This finding again points towards a water replacement stabilization mechanism. The IgG spray-dried powder prepared from the dialyzed liquid feed showed continued substantial aggregation on dry storage at 25°C. This was substantially less in the non-dialyzed, sorbitol-containing spray-dried powder. Addition of trehalose to both dialyzed and non-dialyzed system produced substantial improvement in storage stability and reduction in aggregate formation in storage. The quantitative stabilizing effect of the trehalose was only slightly higher than that of the sorbitol. Taken together, these results indicate that both the sorbitol and trehalose stabilize the IgG primarily by a water replacement mechanism rather than by glassy immobilization. The relevance of this work is its questioning of the importance of the usually considered dominance of glassy stabilization of protein in dried systems of high glass transition temperature, such as trehalose. The low glass transition temperature sorbitol produces almost equal process and storage stability in this case.

Abstract: The objectives of the present study were to investigate the effects of Aquatraining of horses (aqua-treadmill exercise; treadmill manufactured by Equitech – L.u.S. Equipment, Warendorf, Germany) on selected blood parameters [lactic acid concentration (mmol/l), haemoglobin content (g/l)] and on heart-rate variability (HRV) [heart rate (beats per min; b.p.m.), standard deviation of all NN-intervals (SDNN; ms), normalized power of the low and high frequency band (LFnorm, Hfnorm; au), % recurrence, % determinism and ratio(corr)]. Seven horses performed six exercise tests with different work loads (walking (x = 1.56 +/- 0.08 m/s) and trotting (x = 2.9 +/- 0.13 m/s): dry, water above the carpus and water above the elbow). The standardized test-protocol was: 5 min warm-up at walk while the water was pumped in, followed by the 20-min exercise period at walk or trot, followed by a 5-min walk while pumping out the water. Blood samples were taken prior to each test at rest in the stable, as well as exactly 5 min after the end of the 20-min exercise period. Electrocardiograms were recorded during rest and the 20-min exercise period. Compared to rest, neither the chosen velocities, the two water levels, nor the dry tests led to a significant increase of the lactic acid concentration in any horse. The haemoglobin content showed a significant increase as a result of exercise. Significant differences could be found between the heart rates at rest and the six exercise tests and between the mean of the levels 'walking' and the mean of the levels 'trotting'. An exercise-induced change of HRV was characterized by a decreasing SDNN, a significantly higher LFnorm (sympathetic influence) combined with a significantly lower HF(norm) power (parasympathetic activity) and a rising degree of order (significantly higher % determinism and nearly unchanged % recurrence) and stability (significantly rising ratio(corr)) of the recurrence plot. In conclusion, the used training-protocol for aqua-treadmill exercises only represents a medium-sized aerobic work load for horses, but the different levels of burden were indicated especially by changes in HRV.

Abstract: Temperature-dependent EPR and temperature-jump measurements have been carried out, in order to examine the high-spin to low-spin transition of aquomethemogobin (pH 6.0). Relaxation rates and equilibrium constants could be determined as a function of temperature. As a reaction mechanism for the high-spin to low-spin transition, the binding of N epsilon of His E7 to the heme iron had been proposed; the same mechanism had been suggested for the ms-effect, found in temperature-jump experiments on aquomethemoglobin. A comparison of the thermodynamic quantities, deduced form the measurements in this paper, gives evidence that indeed the same reaction is investigated in both cases. Our results and most of the findings of earlier studies on the spin-state transitions of aquomethemoglobin, using susceptibility, optical, or EPR measurements, can be explained by the transition of methemoglobin with H2O as ligand (with high-spin state at all temperatures) and methemoglobin with ligand N epsilon of His E7 (with a low-spin ground state). Thermal fluctuations of large amplitude have to be postulated for the reaction to take place, so this reaction may be understood as a probe for the study of protein dynamics.

Abstract: REASONS FOR PERFORMING THE STUDY: As there are no reports on the real workload of horses that jump fences, this study was undertaken in riding-school horses. OBJECTIVE: To compare the workload of horses jumping a course of fences with that of horses cantering over the same course at the same average speed without jumping fences. The workload variables included heart rate (HR), packed cell volume (PCV), acid-base balance (venous pH, pCO2, HCO3-) and blood lactate (LA), glucose, total protein and electrolyte concentrations. METHODS: Eight healthy riding-school horses performed test A (a course of approximately 700 m with 12 jumps from 0.8-1.0 m high at an average speed of approximately 350 m/min) and test B (same course at the same speed, but without the rails) in a crossover study with at least 4 h between the 2 tests. Before each test the horses were fitted with a heart rate meter (Polar Electro). Blood samples were taken from the jugular vein at rest prior to the test, after warm-up before starting the course, immediately after the course and after recovery. All samples were analysed immediately. RESULTS: The mean +/- s.d maximal HR (beats/min) during the course (184 +/- 17 and 156 +/- 21, respectively) and the mean HR after recovery (75 +/- 6 and 63 +/- 7, respectively) were significantly higher in test A compared to test B (P = 0.001 and P = 0.007 respectively). The mean LA concentrations after the course and after recovery (mmol/l) were significantly higher in test A (3.6 +/- 2.7 and 1.0 +/- 0.9, respectively) compared to test B (0.9 +/- 0.5 and 0.3 +/- 0.1, respectively), (P = 0.016 and P = 0.048 respectively). The mean PCV (I/l) after the course and after recovery was also significantly different between tests A (0.48 +/- 0.04 and 0.39 +/- 0.03, respectively) and B (0.42 +/- 0.04 and 0.36 +/- 0.03, respectively) (P<0.01). The mean pH and the mean HCO3- (mmol/l) after the course were significantly lower in test A (7.40 +/- 0.04 and 28.9 +/- 1.4, respectively) compared to test B (7.45 +/- 0.03 and 30.4 +/- 2.3, respectively) (P<0.05). CONCLUSIONS: This study indicates that in riding-school horses jumping fences, even at a low level competition, provokes a significant workload compared to cantering the same distance and speed without fences. POTENTIAL RELEVANCE: This study makes it clear that the extra workload of jumping fences should be taken into account in the training programmes of jumping horses. Further research with more experienced horses jumping higher fences will reveal the workload for top-level jumping horses.

Abstract: We examined the effects of participation in each of 3 modifications of Day 2 of a 3-day-event on blood and serum variables indicative of hydration, acid:base status and electrolyte homeostasis of horses. Three groups of horses – 8 European (E) horses and 2 groups each of 9 North American horses performed identical Days 1 (dressage) and 3 (stadium jumping) of a 3-day-event. E horses and one group of the North American horses (TD) performed modifications of Day 2 of a 1 Star 3-day-event and the other group of North American horses (HT) performed a Horse Trial on Day 2. Jugular venous blood was collected from each horse on the morning of Day 2 before any warm-up activity, between 4 min 55 s and 5 min 15 s after Phase D and the following morning. Eight E horses, 5 TD horses and 8 HT horses completed the trials. There were few significant differences in acid:base or serum biochemistry variables detected among horses performing either 2 variations of the Speed and Endurance day of a 1 Star 3-day-event, or a conventional Horse Trial. Failure to detect differences among groups may have been related to the low statistical power associated with the small number of horses, especially in the TD group, variation in quality of horses among groups and the different times of the day at which the E horses competed. Differences detected among time points were usually common to all groups and demonstrated metabolic acidosis with a compensatory respiratory alkalosis, a reduction in total body water and cation content, and hypocalcaemia. Importantly, horses of all groups did not replenish cation, chloride, and calcium deficits after 14-18 h of recovery.