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Item Bovine dairy complex lipids improve in vitro measures of small intestinal epithelial barrier integrity(PLOS, 2018-01-05) Anderson RC; MacGibbon AKH; Haggarty N; Armstrong KM; Roy NC; Brandner JMAppropriate intestinal barrier maturation is essential for absorbing nutrients and preventing pathogens and toxins from entering the body. Compared to breast-fed infants, formula-fed infants are more susceptible to barrier dysfunction-associated illnesses. In infant formula dairy lipids are usually replaced with plant lipids. We hypothesised that dairy complex lipids improve in vitro intestinal epithelial barrier integrity. We tested milkfat high in conjugated linoleic acid, beta serum (SureStart™Lipid100), beta serum concentrate (BSC) and a ganglioside-rich fraction (G600). Using Caco-2 cells as a model of the human small intestinal epithelium, we analysed the effects of the ingredients on trans-epithelial electrical resistance (TEER), mannitol flux, and tight junction protein co-localisation. BSC induced a dose-dependent improvement in TEER across unchallenged cell layers, maintained the co-localisation of tight junction proteins in TNFα-challenged cells with increased permeability, and mitigated the TEER-reducing effects of lipopolysaccharide (LPS). G600 also increased TEER across healthy and LPS-challenged cells, but it did not alter the co-location of tight junction proteins in TNFα-challenged cells. SureStart™Lipid100 had similar TEER-increasing effects to BSC when added at twice the concentration (similar lipid concentration). Ultimately, this research aims to contribute to the development of infant formulas supplemented with dairy complex lipids that support infant intestinal barrier maturation.Item Development and validation of an LC-MS/MS method for the quantification of oral-sugar probes in plasma to test small intestinal permeability and absorptive capacity in the domestic cat (Felis catus)(Elsevier BV, 2024-07-15) Patterson K; Fraser K; Bernstein D; Bermingham EN; Weidgraaf K; Kate Shoveller A; Thomas DA novel method for quantifying the concentration of lactulose, rhamnose, xylose, and 3-O-methylglucose (3-OMG) in cat plasma using liquid chromatography-mass spectrometry (LC-MS) was developed. Domestic male cats (n = 13) were orally dosed with a solution containing the four sugars to test the permeability and absorptive capacity of their intestinal barrier. Plasma samples were taken 3 h later and were prepared with acetonitrile (ACN), dried under N2, and reconstituted in 90 % ACN with 1 mM ammonium formate. Stable isotope labelled 13C standards for each analyte were used as internal standards. Chromatographic separation was conducted using a Phenomenex Luna NH2 column with a gradient elution system of deionized water and 90 % ACN with 1 mM ammonium formate at 300 µL/min for 13 min total analysis time. Recovery trials were conducted in triplicate over three days with RSD values (%) for each day ranging from 1.2 to 1.4 for lactulose, 5.4 - 6.0 for rhamnose, 3.3 - 5.5 for xylose, and 2.6 - 5.6 for 3-OMG. Inter-day variations for each analyte were not different (p > 0.05). Limit of detection and quantification were 0.2 and 0.7 µg/mL for lactulose, 0.8 and 2.4 µg/mL for rhamnose, 0.6 and 1.8 µg/mL for xylose, and 0.3 and 1.1 µg/mL for 3-OMG, respectively. Plasma sugar concentrations recovered from cats were above the limit of quantification and below the highest calibration standard, validating the use of this method to test intestinal permeability and absorptive capacity in cats.Item A Simple, Robust, and Convenient HPLC Assay for Urinary Lactulose and Mannitol in the Dual Sugar Absorption Test(MDPI (Basel, Switzerland), 2022-05) Sequeira IR; Kruger MC; Hurst RD; Lentle RG; Locatelli MBACKGROUND: Heterogeneous laborious analytical methodologies for the determination of urinary lactulose and mannitol limit their utility in intestinal permeability testing. METHODS: We developed an assay using a Shimadzu HPLC system, an Aminex HPX87C column, and refractive index detection. The test was calibrated using a series of dilutions from standard stock solutions of lactulose and mannitol 'spiked' into urine samples. The utility to quantify urinary excretion during the dual sugar absorption test over 6 h was also determined. RESULTS: Lactulose and mannitol were eluted isocratically at 5.7 and 10.1 min, respectively, with water as a mobile phase at a flow rate of 0.3 mL min-1, 858 psi, 60 °C. The calibration curves for both sugars were linear up to 500 µg mL-1 with a limit of detection in standard solutions at 4 µg mL-1 and in 'spiked' urine samples at 15 µg mL-1. The intra-assay and inter-assay CVs were between 2.0-5.1% and 2.0-5.1% for lactulose and 2.5-4.4% and 2.8-3.9% for mannitol. The urinary profiles of the 6 h absorption of lactulose and mannitol showed similar peak-retention times to standard solutions and were well-resolved at 5.9 and 10.4 min, respectively. CONCLUSIONS: The assay was easy to automate, using commonly available equipment and convenient requiring no prior laborious sample derivatization. The simplicity, reproducibility, and robustness of this assay facilitates its use in routine clinical settings for the quantification of intestinal permeability.Item The modelling of caking in bulk lactose : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Process and Environmental Technology at Massey University(Massey University, 1997) Bronlund, JohnCaking during storage is a serious problem for manufacturers of bulk lactose. This study was carried out to investigate the causes of caking and identify solutions as to how such problems can be eliminated. The mechanisms for caking in crystalline lactose powders were identified. Liquid bridging between adjacent particles was shown to occur in high relative humidity environments (>80% RH). These liquid bridges could form crystalline solid bridges if the material was subsequently dried out. The potential mechanism of amorphous lactose flow and bridging in conditions where the glass transition temperature is exceeded was shown to be insignificant in predominantly crystalline lactose powders (<5% amorphous lactose). The presence of amorphous lactose is still important as the amorphous matrix acts as a sink of moisture, which can be released upon crystallisation. This increases the moisture available in the system which can contribute to caking by the liquid bridging mechanism. Both of these mechanisms involve changes in the local temperature and moisture conditions within the bulk powder. Such changes were known to be caused by moisture migration under the influence of a temperature gradient. A model which describes the transport of moisture in one dimension as a result of temperature gradients was developed and validated. The microscopic scale processes of liquid bridging and amorphous lactose moisture relations were included into this model. The model predictions agreed well with experimental trials for completely crystalline lactose powders. Comparison of model predictions for the case where amorphous lactose was present on the surface of the particles showed some inadequacies exist in the model. These were the rate of amorphous lactose crystallisation and the assumption of instantaneous equilibrium between the crystallising amorphous matrix and the air present in the interstices of the bulk lactose. Using the model it was shown that for expected storage conditions, the product should be stored with a water activity below 0.57 aw if no amorphous lactose is present and below 0.25 aw if it is present. If these prescribed limits are met then the goal of producing caking free lactose powders can be achieved.