Patent Application
20080058419
The setup of the experiments described below is to assess the influence of DMG on:
To examine all the effects associated with the supplementation of dimethylglycine in fowl feed, a test was setup with broilers as model species. In this slaughter test with 64 broiler hens, 14-day old chickens were bred for 26 days, i.e. till the age of 40 days. At the beginning of the test the broilers were ad random divided in 16 groups of 4 specimens. The birds were fed with one type of feed from the first day on till the end of the test period. The type of feed was attributed on a random basis to the different housings. The animals were setup with a colored ring around their leg for identification purposes within each housing. The control feed was based on a commercial broiler crumble enriched with 5% corn oil to increase the oxidative stress in the feed. Further 1% celite was mixed into the feed as external marker to allow the determination of the apparent nutrient digestibility. The feed had the following composition: 87.65% dry matter, 6.13% ash, 18.05% crude protein, 9.13% crude fat, 4.32% crude fibre and 50.01% other carbohydrates (including 89% starch and 2% sugar). It has a (calculated) metabolizable energy value of about 13.4 MJ/kg.
In the second type of feed 167 mg of DMG was added per kilogram to the same composition of feed as described for the control feed. Over the entire test period, the broilers consumed on average about 25 mg DMG per bird per day. The housing of the broilers consisted out of circular housings with an open roof and bottom and with a total surface of 0.72m2. The material of the fencing was a flexible wire netting with a height of 1 m and mazes of 2×2 cm and thickness of 2 mm. The bottom was strewed with a thick layer of turf with on top a layer of wood-shavings. Feed and water were ad libitum available at all times. The average environmental temperature was maintained at 15° C. to increase the occurrence of ascites (Shlosberg A., Zadikov I., Bendheim U., Handji V., Berman E. Avian Pathology, 21,369-382 (1992)). A conventional light scheme of 23 hours light:1 hour dark was applied Each bird was blood sampled and weighted 3 times, namely at days 1, 15 and 26. The hematocrite concentration was immediately determined by means of an ultracentrifugation on a sub sample of each blood sample. Next, the blood samples were subjected to a centrifugation and the blood plasma was kept at −20° C. On this plasma the concentrations of the plasma metabolites: triglycerines and free fatty acid (NEFA: non-esterified fatty acids) were determined.
The daily growth during the two periods (between the first and fifteenth day and between the sixteenth and twentysixth day) was calculated for each surviving animal. The feed uptake for each housing was also measured during both periods. The commercial feed conversion rate was calculated for each of the housings for both periods (FCRI and FCR II) and for the total growth period. This commercial feed conversion rate was calculated by dividing the total feed consumption by the weight gain of the birds which stayed alive during the entire experiment, including also the feed consumption of the birds that died. To determine only the non-therapeutic effect of DMG on the feed conversion rate, the actual feed conversion rate of the living birds was also calculated more particularly on the basis of weight gain and the feed consumed by these living birds (based on the weight gain of the birds that died, the amount of feed consumed by these birds was detracted from the total feed consumption).
At the start of the second period a sample of 100 g of manure was collected. The coefficients for the apparent metabolizability of macronutrients and the apparent nitrogen retention was calculated based on the external marker method with the acid insoluble (celite) as external marker
Apparent metabolizability =1−(NF/NV×IV/IF) wherein : NF: percentage of examined nutrient in the faeces sample
NV: percentage of examined nutrient in the feed sample
IV: percentage of the indicator in the feed sample
IF: percentage of the indicator in the faeces sample
To determine the above parameters both the faeces and the feed samples were analysed on the dry matter, crude ash, crude protein, crude fat and crude fibre (AOAC, 1980). The carbohydrates were calculated from the difference between the dry matter content and the rest of the macronutrients as determined by means of the Weende analysis. The content of insoluble ashes (celite) was determined according to the procedure of Atkinson et al. (1984).
Finally there was also an autopsy on all chickens. First, the carcass weight was determined by dissection of the head, the paws, evisceration and the deduction of the average feather weight. Next, the weight of the breast, buttock and thigh muscle were determined. Also the the heart, liver and abdominal fat were measured. Macroscopic lesions related to ascites were visually reviewed and described as indicated by Scheele et al. (2003) (Scheele, C. W., Van Der Klis, J. D., Kwakernaak, C., Buys, N., Decuypere, E., British Poultry Science, 44(3), 484-489 (2003)), these were: accumulation of liquid in the abdomen, a hydropericard and right heart dilatation. The latter was quantified by the ascites heart index (AHI) being the ratio of the the dry weight of the right ventricle to the dry weight of both ventricles after freeze drying.
Ascites was the major cause of death during the test period of 26 days. The mortality and occurrence of ascites were substantially lower in the DMG supplemented group (Table 1 and 2). Next to a massive accumulation of fluid in the abdomen, all these animals showed a distinct right heart ventricle dilatation. At the beginning of the second phase of the experiment the chickens fed with the DMG supplementation showed a higher apparent metabolizability of the dry matter. From the calculated metabolic coefficients for the dry matter and the proteins it can be seen that the ability to metabolise both dry matter and proteins seems to be improved in the DMG supplemented group (Table 3). From the second period on, the DMG supplemented group also showed a significantly improved feed conversion and also an increased growth rate. The total growth rate was however substantially not affected so that the improved feed conversion has to be explained by a lower feed consumption.
In the above experiment, the actual occurrence of the broiler ascites syndrome of 4.7% (Maxwell H. M., Robertson G. W. British Poultry Science 39,203-215 (1998)) was significantly increased by the setup of the test. The inventors could clearly show that there was a significant difference in the occurrence of ascites in respectively the DMG supplemented and control group of animals. Hence, it could be concluded that DMG has a protective effect on the pathogenesis of ascites. This effect was also confirmed in the increased hematocrite values for the DMG supplied group of animals. The direct proof of the positive effect of DMG on the ascites condition was found during the autopsy tests on the hart of the chickens. For all animals the AHI (Ascites Heart Index)) was calculated as the ration of the dry weight of the right heart ventricle to the dry weight of both heart ventricles. This parameter gives a clear indication on the occurrence of right heart hypertrophy (symptom of ascites syndrome). Significantly higher AHI values were found for the group of chickens who weren't supplemented with DMG.
Not only was the ascites syndrome positively influenced by DMG, it was further found that the DMG supplementation led to a reduced feed conversion rate, especially during the second growing period. The commercial feed conversion rate, which included the therapeutic effect of DMG on ascites and which is very important from a practical point of view was clearly improved. However, also the actual feed conversion rate, calculated only on the basis of the weight gain and of the feed consumed by the birds which stayed alive, was clearly improved. (The same actual feed conversion rate can also be calculated on the basis of total weight gain of all animals, including the birds that died during the trial). This proves that DMG has also a non-therapeutic effect on the feed conversion rate. This non-therapeutic effect was moreover coupled to an increase of dry matter and protein metabolizability for the chickens fed with the DMG supplemented diet.
In the blood panel of the animals, substantial differences in the concentration of triglycerides and significant differences in the concentration of free fatty acids (NEFA) were detected. The decrease of free fatty acids, with the DMG supplemented group, can be explained by either an increased extraction of these products from the blood or by the fact that less NEFA were mobilized from the fat reserves of the animals. Since DMG also gives rise to an increase in apparent metabolizability of the feed, it is most likely that the reduced values for the NEFA were due to the fact that there were less NEFA mobilized from the fat reserves of the chickens. This conclusion can be further supported by the high levels of triglycerides detected in the blood samples of the DMG supplemented group. This leads to the conclusion that DMG can be important as support for the energy metabolism and the reduction of metabolic stress.
In summary, it was found that the supplementation of 0.001-0.5 wt. % (based on the feed) of the glycine compound, more particularly of DMG or its salts, has a beneficial effect on a disease in fowl called ascites. The DMG supplementation leads to an increase of the hematocrite level so that the organism of the bird can deal more effectively with limited oxygen supply to the tissues. Moreover the DMG also plays a role in the reduction of the free fatty acid content in the blood stream which can lead due to the presence of unsaturated bonds in the molecular structure to oxidative stress leading to the death of the animal. This property is important in the modern feed formulation technology which uses more and more vegetal fats to supplement the feed. Further DMG has a distinct influence on both the commercial and the actual feed conversion, an important economic parameter in the cultivation of fowl in general. This feed conversion is directly linked to the effect that a DMG supplementation has on the apparent metabolizability of both dry matter and proteins. In general it was found that DMG has a positive influence on the occurrence of ascites in fowl and on the bird's energy metabolism.
| Filing Document | Filing Date | Country | Kind | 371c Date |
|---|---|---|---|---|
| PCT/EP06/60926 | 3/21/2006 | WO | 00 | 9/20/2006 |
