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    The use of the Ussing chamber system to investigate iron absorption by the duodenum, jejunum and ileum in the mouse : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Physiology at Massey University, Palmerston North, New Zealand
    (Massey University, 2002) Edmunds, Shelley Jane
    Iron deficiency anaemia is found in approximately 30% of the worlds population and is particularly prevalent in developing countries. The majority of these deficiencies are due to insufficient absorption of iron from the diet. Iron is absorbed primarily by the proximal small intestine, however, there is evidence for a gradient of absorption along the full length of the small intestine. In 1951 Ussing and Zerahn developed a bicameral method for studying iron transport by in vitro epithelia. This method has been used previously to investigate iron transport mechanisms in the proximal small intestine. In the present study Ussing chambers were used to investigate iron absorption by the full length of the mouse small intestine. Consistently high levels of iron were removed from the mucosal compartment by all regions of the small intestine. This iron removal was due to the physiological actions of the tissue and was not caused by iron adhering to the interior of the Ussing chamber apparatus. There was no change in iron uptake when large intestine or caecum was used in place of small intestine. Ferrous gluconate was chosen as the reference test chemical as it is a readily bioavailable form of iron which has been used previously to investigate iron absorption with the Ussing chamber model. There was a consistently high level of iron uptake when 27.9 mg/L or 9.3 mg/L was added to the mucosal compartment, with no significant differences between results for either concentration. When 9.15 mg/L manganese sulphate was combined with 9.3 mg/L ferrous gluconate in the mucosal compartment, iron removal was significantly lower in the proximal than the mid small intestine. This was presumably due to competition between the iron and the manganese for transport by the DCT1 protein. When 200 mg/L calcium chloride and 9.3 mg/L ferrous gluconate were added to the mucosal compartment, there was no significant difference to results compared to ferrous gluconate alone. The addition of glucose to the intestinal lumen has been shown previously to increase the passive transport of solutes across the intestinal mucosa. However, in the present experiments when glucose was added to the mucosal Ringer's solution in place of mannitol there was a significant decrease in iron removed from the mucosal compartment by all intestinal regions. There was evidence that the gluconate portion of ferrous gluconate increased iron absorption in the distal small intestine. This was supported by a significant decrease in iron uptake by the distal small intestine when ferrous sulphate was used in place of ferrous gluconate. Ferric chloride was unsuitable for use in this system as it precipitated out of the Ringer's solution. Histological examination of jejunal samples after a typical Ussing chamber experiment found there was no damage to the tissue and the epithelial layer remained intact. There were significant levels of iron found in both the intestinal tissue and secreted mucus for all intestinal segments. The binding of iron to secreted mucus appears to involve a significant proportion of iron and should be measured in all future Ussing chamber studies.
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    Pre- and pro-biotics may improve mineral absorption and retention in the growing male rat : a thesis submitted in partial fulfilment of the requirements for the degree of Master of Science in Nutritional Science at Massey University, Turitea, Palmerston North, New Zealand
    (Massey University, 2005) Fear, Alison Lindsay
    Probiotics are bacteria, which reside in the large intestine and concur beneficial health effects on their host. Their abundance can be selectively-stimulated by prebiotics, such as fructo-oligosaccharide (FOS); prebiotics are oligosaccharides, which are not digested in the small intestine, but pass into the large intestine where they are fermented into short-chain fatty acids. Several studies have suggested that prebiotics may improve mineral absorption. This study aimed to determine the effects of pro- and pre-biotic supplementation on mineral absorption and bone quality in growing male rats. Sixty three-week old male Sprague-Dawley rats were randomised into five groups and fed either a high-calcium milk powder (HCMP) with or without a probiotic added (groups were subsequently named HCMP - and HCMP + respectively), or HCMP and vitamin K with or without the probiotic (HCMPK - and HCMPK +), or the HCMP with FOS replacing the sucrose in other diets, and the probiotic (the dietary group was named FOS). Animals were maintained on diets for 10 weeks. Balance studies were carried out during weeks 3 - 4 and 8 - 9 of the study. The earlier balance study suggested that dietary interventions may affect mineral absorption. The latter balance study, however, showed no discernable differences between groups. Several reasons were postulated for this. Active-absorption may have been down-regulated as a result of long-term supplementation, or an increased abundance of probiotics could cause an elevation of nutritional demands. Alternatively, supplementation may not prove beneficial once animals had passed their period of peak absorption. Bone resorption and formation did not appear to have been altered as a result of dietary intervention, when measured after 10-weeks. Bone mineral density and content, calcium, magnesium, zinc and ash contents and bone biomechanical testing also showed no significant differences between dietary groups. Further research is required to determine whether results obtained were due to long-term supplementation and / or due to the joint-supplementation of pre- and pro-biotics.
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    Adherence interactions between milk proteins and human intestinal surface layer components : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy, Massey University, Palmerston North, New Zealand
    (Massey University, 2016) Schmidmeier, Christiane
    Recent research suggests a number of food-derived proteins may be used as orally delivered functional components. The native structure is often vital to their activity and requires protection during the digestive process. Nutrient vehicles are used as protective envelopes and as a mechanism for targeting specific sites of activity, e.g. the small intestine. This study evaluated molecules which adhere to one or more in vitro models of three human intestinal surface layers. Successful candidates could then be incorporated into nutrient vehicles, promoting adhesion to the surface layers and resulting in prolonged retention of the active ingredient at the site of action or absorption. To identify molecules that adhere to models of the intestinal surface, an adhesion protocol was developed to screen the proteome of whole milk, skim milk and whey for candidate proteins. Molecules adhering to model layers of the human gastrointestinal tract (intestinal epithelial cells, mucin or bacteria with the propensity to form a biofilm) were screened by SDS-PAGE analysis and identified by mass spectrometry and Western blot. The binding behaviour of selected proteins was further investigated by flow cytometry. The combined results showed that milk and whey proteins exhibit different binding affinities to the models of individual surface layers. a-Lactalbumin was found to adhere to a model of the intestinal epithelial cells, while ß-lactoglobulin showed binding to the protective mucin layer. Lactoferrin and various components of immunoglobulins showed highest binding affinity to bacteria. Finally, IgM appeared to adhere to all three tested model layers of the human gastrointestinal surface. Least binding was observed to the intestinal epithelial cells in culture. The validity of the developed adhesion protocol was demonstrated by replicating adhesion of immune-related proteins, lactoferrin and immunoglobulins, to bacterial cells. This work reveals new important characteristics of milk-derived proteins in their ability to adhere to models of the gastrointestinal surface. These may be further utilised in site-specific targeting of functional foods.
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    Factors influencing fat digestion in poultry : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Poultry Nutrition at Massey University, Palmerston North, New Zealand
    (Massey University, 2012) Tancharoenrat, Piyamas
    The first experiment in this thesis was conducted to understand the digestion of fat along the gastrointestinal tract and ileal endogenous fat losses. The second, third and fourth experiments investigated the factors influencing fat digestion in broilers, including age of birds, cereal type, fat source and dietary calcium concentrations. In the fifth experiment, influence of unsaturated to saturated fatty acid ratio (U:S ratio) on fat digestion was examined. The first experiment reported in Chapter 4 showed that jejunum is the major intestinal site where majority of fat and fatty acids is digested and absorbed. Long chain fatty acids showed lower digestibility than short chain fatty acids, and unsaturated fatty acids were better digested than saturated fatty acids. The fatty acid profile of ileal endogenous fat was remarkably similar to that of the bile, suggesting that the reabsorption of fat and fatty acids in bile was incomplete. Data reported in Chapter 5 showed that the apparent metabolisable energy (AME) and total tract digestibility of fats was influenced by the age of broilers. The AME of fat was markedly lower during the first week, increased rapidly during second week and then remained constant thereafter. Total tract fat digestibility was poor during the first week and then increased until the third week of age. No further improvement was observed after the third week. The AME and fat digestibility of soybean oil, poultry fat and palm oil were determined to be higher than those of tallow. Blending of tallow and soybean oil resulted in AME and fat digestibility estimates higher than the arithmetic averages of tallow and soybean oil. The study reported in Chapter 6 showed that the supplementation of tallow in wheat- and maize-based diets resulted in lower weight gain than that of soybean oil, but fat source had no effect on the weight gain of broilers fed sorghum-based diets. Broilers fed soybean oil supplemented diets had lower feed per gain, higher total tract retention and ileal digestibility of fat compared to those fed tallow supplemented diets. In addition, supplementation of xylanase in wheat-based diets resulted in improved weight gain and feed efficiency of broiler starters irrespective of the fat source. Xylanase supplementation increased the AME of tallow supplemented diets, but had no effect on soybean oil supplemented diets. Data reported in Chapter 7 indicated that high dietary calcium concentrations had negative impact on broiler performance, irrespective of tallow inclusion levels. High calcium concentrations resulted in higher excreta soap and, lowered the total tract retention of fat, calcium and phosphorus. Lower calcium concentrations resulted in higher ileal digestibility of fat, nitrogen and phosphorus. Data from the final experiment (Chapter 8) showed that the U:S ratio influenced the performance of broilers during the starter period (1 to 21 day), but had no effect on the performance over the whole trial period (1 to 35 day). Increasing the U:S ratio decreased the AME of diet and increased the total tract retention of fat. A positive linear correlation between U:S ratio and the AME of fat blends was observed, with increasing U:S ratios improving the AME of fat blends. In conclusion, the research reported in this thesis identified several factors that influence the digestion of fat in poultry. Age of broilers influenced the digestion and absorption of fat, particularly during the first week of age. The findings consistently demonstrated that tallow was more poorly digested than soybean oil. The utilisation of these two fat sources was influenced by the cereal base used in the diets, with the effect of fat source on weight gain differing between cereal types. High dietary calcium concentrations were detrimental to the digestibility of nutrients, especially of fat. A finding of practical interest was that digestion and absorption of animal fats, containing high concentrations of saturated fatty acids, can be improved by blending with soybean oil to increase the U:S ratio.