Ross × Ross 708 eggs were injected with commercial diluent containing supplemental L-carnitine at 8, 16, or 32 mg/100 μL concentrations using an automated multi-egg injector. After hatching, 1,080 male and female broiler chicks were distributed into 90 pens with chicks at each of the injected concentrations receiving feed that was or was not supplemented with 50 ppm of L-carnitine. Treatments did not affect incubation time or hatchability of fertilized eggs. Birds fed supplemental L-carnitine and injected in ovo with L-carnitine had lower body weight and ate less feed. The same birds exhibited a reduction in feed conversion compared to birds that did not receive supplemental dietary L-carnitine. Absolute breast weight was reduced in birds given L-carnitine in ovo and in the feed. Broiler diets containing 50 ppm L-carnitine appeared to be slightly toxic if provided with 8, 16, or 32 mg/100 μL of L-carnitine administered via in ovo injection.

ABSTRACT This thesis research involved investigations into the effects of inflammation on maturation of L-carnitine homeostasis in developing rat neonates. The overall hypothesis was an inflammatory stimulus will alter the ontogeny of L-carnitine homeostasis pathways and this depends upon when the inflammatory stimulus occurs in postnatal development. The objective was to investigate the potential effect of inflammation on carnitine transporter expression in different age groups of neonates and evaluation of effect of inflammation on ontogeny and activity of enzymes involved in carnitine biosynthesis and whether this differs depending upon when in postnatal development the inflammatory stimulus occurs. Rat pups at postnatal day 3, 7, and 14 received an intraperitoneal injection of lipopolysaccharide (LPS) at a dose known to cause a febrile reaction in rat neonates. L-Carnitine homeostasis pathways underwent significant ontogenesis during postnatal development in the rat. LPS administration caused a significant decrease in free L-carnitine levels in serum and heart tissue and a decrease in mRNA expression levels of the high affinity carnitine transporter, Octn2, in kidney, heart and intestine at all postnatal ages. Furthermore, significant decreases in mRNA expression levels of key enzymes involved in carnitine biosynthesis was observed, while an increase in carnitine palmitoyltransferase mRNA levels were observed at all postnatal ages. Reductions in butyrobetaine hydroxylase mRNA expression were paralleled by reductions in enzyme activity only at postnatal day 3 and 7. Heart creatine phosphate levels were deceased significantly in LPS treated groups in all postnatal ages; however, ADP and ATP levels were unaffected. Collectively, this research provided experimental evidence for a significant effect of inflammation on changes in L-carnitine homeostasis maturation in early neonatal stages. The maturation of physiological processes may be altered by external factors in early postnatal life.

Master of Science / Department of Animal Sciences and Industry / Barry J. Bradford / Dairy cows undergo many homeorhetic adaptations during the transition to lactation. Although many of the physiological processes - including increased lipolysis and postpartum inflammation - are adaptive, exaggerated responses can contribute to metabolic disease and reduced milk production. L-carnitine has been shown to increase hepatic oxidation of fatty acids and reduce hepatic lipid accumulation in early lactation cows; however, L-carnitine is degraded in the rumen. An experiment using 4 ruminally-cannulated Holstein heifers in a split plot design demonstrated that the relative bioavailability of L-carnitine was greater when delivered abomasally than ruminally. There was a dose × route interaction and a route effect for increases in plasma carnitine above baseline, with increases above baseline being greater across all dose levels (1, 3, and 6 g L-carnitine/d) when infused abomasally compared to ruminally. A second experiment used 56 lactating Holstein cows in a randomized complete block design to evaluate 2 rumen-protected products (40COAT and 60COAT) compared to crystalline L-carnitine at doses targeting 3 and 6 g/d carnitine. Although crystalline and 40COAT were effective in linearly increasing carnitine concentrations, only subtle responses were seen for the 60COAT, which were less than that for crystalline carnitine in plasma, milk, and urine. Ineffectiveness of rumen-protected products to increase carnitine concentrations beyond crystalline may have been due to over-encapsulation that hindered liberation of the carnitine and its absorption in the small intestine. Although L-carnitine has the potential to reduce postpartum hepatic lipidosis, effective rumen protection of L-carnitine while maintaining intestinal availability needs further investigation. Plant polyphenols have anti-inflammatory properties and when administered during the transition period, have been shown to increase milk production. An experiment used 122 multiparous Holstein cows in a randomized block design to determine the effect of short term (5-d; SBE5) and long term (60-d; SBE60) administration of Scutellaria baicalensis extract (SBE)on whole-lactation milk yield, 120-d milk component yield, and early lactation milk markers of inflammation. Whole-lactation milk yield was increased for SBE60 compared to control, but was not different for SBE5 compared to control. Greater total pellet intake, milk lactose yield, and reduced SCC during wk 1-9 for SBE60 compared to control, all could have contributed to the observed sustained increase in milk yield. Milk production parameters were not different for SBE5 compared to control. No treatment effects were observed for BCS or milk markers of inflammation (haptoglobin) and metabolic function (β-hydroxybutyrate). Overall, long term administration of S. baicalensis effectively increased milk production, however the mechanism by which this was achieved is unknown. Although routes of administration to effectively achieve their physiological responses were different between L-carnitine (abomasal delivery) and SBE (feeding), both bioactive nutrients can improve the metabolic function of early lactation dairy cows.

In the first phase of the current study, metabolic combinatorial solutions were in ovo administered in broiler eggs on Day 18 of incubation to investigate their effects in broiler chick tissue nutrient profiles until Day 10 of posthatch growout. In the second phase, the effects of in ovo injection of L-carnitine on Day 18 of incubation in broiler eggs were examined. The treatment solutions used in these studies were considered to play significant roles in various biochemical pathways, and were hence tested to determine whether they could potentiate the physiological growth and development of the broiler embryos and posthatch chicks. The in ovo injection of treatment solutions in both trials did not produce any significant increase in performance or slaughter yield in broiler chicks. However, positive trends were determined for rate of hatch and tissue nutrient profiles, which implied that the in ovo administration of nutrient supplements may be supportive of embryonic development and posthatch growout performance.

This thesis (Application of L-carnitine to the peritoneal cavity of rat) deals with the effect of L-carnitine on the inflammatory response of the organism. L-carnitine is a common part of the body's physiological functions. It is part of the lipid metabolism in the mammalian organism. Its effect helps to transfer fatty acids from the cytosol to the mitochondria. The literary part deals with the current state of L-carnitine research and its effects in the body. It also deals with methods of applying active substances to or-ganisms and blood elements, namely leukocytes and their functions associated with inflammatory reaction. The experimental section deals with the influence of L-carnitine on the course of inflammatory reaction in the peritoneal cavity. It also deals with the determination of the dynamics of the inflammatory response, which is evalu-ated using absolute and differential leukocyte counts at two time points.

This set of experiments evaluated the physiological responses of intensively managed ostriches to L-carnitine. In experiment 1, 32 female and 16 male Zimbabwean Blue Neck and South African Black Neck breeders (n=48 of each sub-species; eight years old), were investigated in 16 breeder units of two females and one male (Trio), in a completely randomised design within four treatments and four replicates over an 8-month period during the breeding season. The same basal diet was fed supplemented with 0 (T0, control), 125 (T125), 250 (T250) or 600 (T600) mg/kg L-carnitine. T600 improved the egg production percentage, egg fertility percentage and the hatchability of set eggs for Black-Necks and Blue-Necks, respectively, and the hatchability of fertile eggs in Black Necks. L-carnitine did not affect egg shape index, defective eggs, egg weight, embryonic and post-hatch mortality. In experiment 2, 12 Black Neck males (5.5 years old) were allocated to three treatments (T0, T250 and T500) and four replicates. Semen samples were collected once a month over three months. L-carnitine had a significant effect on semen volume, sperm motility, live sperm percentage and sperm count, but had no significant effect on abnormal sperm percentage. In experiment 3, 32 day-old Black Neck ostrich chicks were allocated to treatments T0, T125, T250 and T600 with four replicates of two chicks. Chicks were vaccinated against inactive Newcastle Disease (ND) vaccine at day 30 as primary, and at day 51 as booster immunisation. ND antibody responses in the sera were monitored over three phases at 51, 70 and 80 days. Anti-NDV antibodies were detected using a modified chicken anti-NDV enzyme-like immunosorbent assay (ELISA). The treatments and the time periods and their interactions influenced ND antibody responses. T125 and T250 had the highest level of ND antibody response compare to the other groups. There were no differences in ND antibody response between T0 and T600 as well as T125 and T250. The highest ND antibody responses were recorded at day 70. Experiment 4 was designed the same as 3, to determine chicks’ growth responses over the 60-day period. Live weight and live weight gain values of T125 and T250 did not differ from those of T0. T600 had the lowest feed conversion ratio (FCR) during the total period. Feed intake (FI) was reduced in the T125 and T600 treatments compared to T0 and T250 over the total period. T125 gave the lowest FI and FCR responses over the total period, whereas there was no difference between T0 and T250. These results suggest that dietary T600 can have a beneficial effect on egg production, fertility and hatchability in the Black and Blue Neck breeders and T250 might improve sperm quality in males. In ostrich chicks T125 and T250 had positive effects on immune responses and T125 can improve the performance by decreasing the FCR. In contrast, the suppressive effect of a high inclusion level (T600) might indicate that ostrich chicks are sensitive to high inclusion levels that could cause adverse effects. / gm2013 / Animal and Wildlife Sciences / Thesis (PhD)--University of Pretoria, 2013. / Unrestricted

The objectives were to: 1) determine if im treatments of Lutalyse expedited the training of sexually inexperienced boars for semen collection and increased spermatozoal output, and 2) determine the effects of dietary L-carnitine supplementation on boar libido, semen quality, sperm production, and maintenance of sperm motility during liquid storage. Experiment 1 utilized lean-type, terminal-line boars (National Pig Development, Roanoke Rapids, NC) (n = 40; 177.4 ± 2.4 d of age and 112.8 ± 2.0 kg body weight) that had not previously experienced natural mating. Boars were individually moved twice weekly for 6 weeks (total of 12 training sessions) to a semen collection room equipped with an artificial sow. Upon entering the semen collection room, boars received in treatments of either deionized water (4 mL, n = 10) or Lutalyse at doses of 5 mg (n = 10), 10 mg (n = 10), or 20 mg (n = 10), and subsequently received a libido score of 1 to 5 (1 = no interest in the artificial sow; 5 = mounting the artificial sow and allowing semen collection). The percentages of boars successfully trained for semen collection during the experimental period were similar (P > 0.05) for controls (20%) and boars receiving 5 mg (30%), 10 mg (20%), or 20 mg (10%) of Lutalyse. Average libido score for boars receiving 10 mg Lutalyse (2.35 ± 0.08) was greater (P < 0.05) than for controls (2.14 ± 0.06). Libido score for the 20 mg treatment group were (1.78 ± 0.06) lower (P < 0.05) compared to the other treatment groups. Characteristics of ejaculates (volume, gel weight, sperm concentration, total spermatozoa) from control boars and boars treated with Lutalyse at doses of 5, 10, or 20 mg were similar (P > 0.05). For Exp. 2, the same group of boars was utilized in two similar trials (Trial 1, 1a, 1b: n = 9 for control and L-carnitine-treated boars; Trial 2, 2a, 2b: n = 10 for control and L-carnitine-treated boars). Boars were fed a fortified, corn and soybean meal-based diet at a rate of 2 kg/d. Boars that were randomly selected for L-carnitine treatment received the same diet mixed with L-carnitine to achieve supplementation of 500 mg/d. For 16 wk, semen was collected weekly via the gloved hand method and was analyzed for gel-free volume, gel weight, sperm concentration, sperm per ejaculate, and characteristics of sperm motility. Time to ejaculation (reaction time), duration of ejaculation, and number of false mounts were also recorded for each collection. Trials 1a and 2a were conducted during weeks 16 and 17 for each respective trial. Boars were collected once on 4 consecutive days, allowed 4 d of rest, and then collected again, to estimate daily spermatozoal production. At the end of 16 wk, a semen sample was also processed and extended in Beltsville Thawing Solution (BTS) to achieve a dilution of 3 x 109 spermatozoa/100 mL-dose for Trials 1b and 2b. The extended semen was stored in plastic bottles at 18°C and motility was evaluated daily for 7 d post collection. L-carnitine supplementation for 16 wk had no effects on semen volume, gel weight, total number of sperm cells per ejaculate, reaction time, or sperm motility (P > 0.1). Boars receiving the L-carnitine-supplemented diet displayed an increase in the number of false mounts before ejaculating and an increase in sperm concentration (P < 0.05) in Trial 2. A treatment by week interaction was detected for sperm concentration in Trial 2 (P < 0.005). Increased sperm concentrations in L-carnitine-treated boars were demonstrated after only one week of feeding the respective diets. Given that the production of a mature sperm cell requires 7 to 8 wk in boars, it is therefore difficult to conclude that differences in sperm concentration were due solely to treatment. Daily spermatozoal production was similar between control boars and boars supplemented with L-carnitine (P > 0.1) for both Trials 1a and 2a. L-carnitine supplementation did not affect percent motility in Trials 1b and 2b or sperm progressive motility in Trial 2b during 7 d storage (P > 0.1). A treatment by day interaction was determined for sperm velocity (P < 0.05) in Trial 2b. L-carnitine supplementation decreased mean sperm velocity significantly after 2 d of storage. Overall, L-carnitine had no beneficial effects on boar libido, semen quality, sperm production, or maintenance of sperm motility during liquid storage. However, Lutalyse increased libido scores, but did not affect the number of boars trained for semen collection or number of spermatozoa ejaculated. / Master of Science

Doctor of Philosophy / Department of Animal Sciences and Industry / Michael D. Tokach / Four experiments were conducted to determine the interactive effects of dietary L-carnitine and ractopamine HCl (ractopamine) on finishing pig growth performance. In analysis of treatments common to all experiments, ractopamine increased (P < 0.01) ADG and G:F compared to pigs not fed ractopamine. Added L-carnitine tended to increase (P < 0.07) ADG and improved (P < 0.01) G:F compared to pigs not fed L-carnitine. Three experiments were conducted to determine the effects of L-carnitine and ractopamine on carcass characteristics and meat quality. In Exp. 1, drip loss decreased (linear, P < 0.04) in pigs fed increasing L-carnitine. In Exp. 2, drip loss decreased (P < 0.04) with increasing L-carnitine when fed with ractopamine. Percentage lean was higher (P < 0.01) for pigs fed ractopamine. In Exp. 3, lean percentage increased (P < 0.03) in pigs fed L-carnitine or ractopamine. Pigs fed L-carnitine tended (P < 0.06) to have decreased drip loss. These results suggest that ractopamine increases carcass leanness and L-carnitine reduces drip loss when fed in combination with ractopamine. Two experiments were conducted to determine the effects of L-carnitine and ractopamine on the metabolic response to handling. Non-gentle handling increased (P < 0.01) lactate and rectal temperature, and decreased pH. In Exp. 1, non-gentle handled pigs fed ractopamine had decreased (P < 0.01) pH and increased temperature and tended (P < 0.09) to have higher lactate than other pigs. In Exp. 2, lactate and temperature changes from immediately post-handling to 1 h post-handling were not different for pigs fed L-carnitine or ractopamine suggesting that L-carnitine did not decrease recovery time of pigs subjected to non-gentle handling or fed ractopamine. These results suggest that pigs fed ractopamine are more susceptible to stress when handled aggressively. Because carnitine did not alleviate the negative effects of handling for pigs fed ractopamine, the improvement in drip loss from feeding carnitine must be due to a different mode of action.

Traumatic injuries to the spinal cord and brachial plexus induce a significant inflammatory response in the nervous tissue with progressive degeneration of neurons and glial cells, and cause considerable physical and mental suffering in affected patients. This thesis investigates the effects of the antioxidants N-acetyl-cysteine (NAC) and acetyl-L- carnitine (ALC) on the survival of motoneurons in the brainstem and spinal cord, the expression of pro-apoptotic and pro-inflammatory cell markers, axonal sprouting and glial cell reactions after spinal hemisection in adult rats. In addition, a novel MRI protocol has been developed to analyse the extent of neuronal degeneration in the spinal cord. Rubrospinal neurons and tibial motoneurons were pre-labelled with the fluorescent tracer Fast Blue one week before cervical C3 or lumbar L5 spinal cord hemisection. The intrathecal treatment with the antioxidants NAC (2.4mg/day) or ALC (0.9 mg/day) was initiated immediately after injury using Alzet2002 osmotic mini pumps. Spinal cord injury increased the expression of apoptotic cell markers BAX and caspase 3, induced significant degeneration of rubrospinal neurons and spinal motoneurons with associated decrease in immunoreactivity for microtubule-associated protein-2 (MAP2) in dendritic branches, synaptophysin in presynaptic boutons and neurofilaments in nerve fibers. Immunostaining for the astroglial marker glial fibrillary acidic protein and microglial markers OX42 and ED1 was markedly increased. Treatment with NAC and ALC attenuated levels of BAX, caspase 3, OX42 and ED1 expression after 2 weeks postoperatively. After 4-8 weeks of continuous intratheca ltreatment, NAC and ALC rescued approximately half of the rubrospinal neurons and spinal motoneurons destined to die, promoted axonal sprouting, restored the density of MAP2 and synaptophysin immunoreactivity and reduced the microglial reaction. However, antioxidant therapy did not affect the reactive astrocytes in the trauma zone. The inflammation modulating properties of ALC were also studied using cultures of human microglial cells. ALC increased the microglial production of interleukin IL-6 and BDNF, thereby possibly mediating the anti-inflammatory and pro-regenerative effects shown in vivo. To study degeneration in the spinal cord following pre-ganglionic and post-ganglionic brachial plexus injuries, adult rat models of ventral root avulsion and peripheral nerve injury were used. A novel MRI protocol was employed and the images were compared to morphological changes found in histological preparations. Ventral root avulsion caused degeneration of dendritic branches and axonal terminals in the spinal cord, followed by significant shrinkage of the ventral horn. Extensive astroglial and microglial reactions were detected in the histological preparations. Peripheral nerve injury reduced the density of dendritic branches but did not cause shrinkage of the ventral horn. Quantitative analysis of MRI images demonstrated changes in the ventral horn following ventral root avulsion only, thus validating the developed MRI technique as a possible tool for the differentiation of pre-ganglionic and post-ganglionic nerve injuries.

Master of Science / Department of Animal Sciences and Industry / Joel DeRouchey / Mike Tokach / Six experiments using 3,862 pigs were conducted to evaluate effects of ractopamine HCl (RAC) feeding programs, dietary L-Carnitine and dried distillers grains with solubles (DDGS) on growth, carcass traits, loin and jowl fat quality of pigs, and energy and protein sources in nursery diets. In Exp. 1 and 2, RAC-fed pigs had greater (P<0.05) ADG, G:F and HCW compared with the control. Within RAC treatments, there were no differences in growth. Pigs fed step-up RAC had increased (P<0.01) percentage lean, fat-free lean index and loin depth but decreased (P<0.01) backfat than the control or constant treatment. In Exp. 2, pigs fed step-up RAC program had greater (P<0.05) ADG and G:F than the constant treatment. Pigs fed constant RAC had greater (P=0.002) carcass yield than controls. There were no overall differences in other carcass traits among treatments. In Exp. 3, dietary L-Carnitine improved (P<0.02) ADG and final BW. A DDGS × L-Carnitine interaction (quadratic, P<0.01) was observed for G:F. Pigs not fed DDGS had similar G:F, but in DDGS diets pigs fed 50 ppm L-Carnitine had worse G:F than those fed 100 ppm. Pigs fed L-Carnitine had greater (P<0.02) HCW compared with those not fed L-Carnitine. Increasing L-Carnitine up to 100 ppm increased HCW (quadratic, P<0.03) and backfat (quadratic, P<0.04), with the maximum response at 50 ppm dietary L-Carnitine. Increasing L-Carnitine increased (linear, P<0.04) purge loss of loin. Feeding DDGS increased (P<0.001) linoleic acid and iodine value of jowl fat compared with feeding no DDGS. However, feeding L-Carnitine did not change jowl fatty acid composition. In Exp. 4, 5 and 6, nursery pigs fed choice white grease (CWG) had improved (P<0.02) G:F than pigs fed a control diet or an alcohol based energy source. Also, pigs fed CWG had greater (P<0.04) ADG in Exp. 4 and 6 and had reduced (P<0.01) ADFI in Exp. 5. The alcohol based energy source improved (P<0.04) ADG and ADFI with no change in G:F in Exp. 4; but did not affect growth in Exp. 5 and 6. In Exp. 6, pigs fed AV-E Digest had equal performance as nursery pigs fed other specialty proteins.