Aliskiren (ALS) is a direct renin inhibitor with low bioavailability and high drug cost. The goal of this study was to increase the bioavailability of ALS through nanoformulation. The optimized formulation was then evaluated in spontaneously hypertensive rats (SHRs). We developed an ALS poly(lactic-co-glycolic) acid nanoparticle (ALS-NP) through the emulsion–diffusion–evaporation method with various solvents, stabilizer concentrations, and centrifugation speeds. SHRs were orally dosed with 30 mg/kg ALS or dose equivalent ALS-NP. Several parameters were assayed in plasma and/or urine at baseline and 24 h post-dose, while pharmacokinetic analysis included serial sampling. The optimum formulation was found with ethyl acetate, a 1.00% w/v didodecyldimethylammonium bromide concentration, and a 10,000 r/min (15,554 g) centrifugation speed. A 168% relative bioavailability was observed as a result of ALS-NP administration along with significant, as determined by Student’s t-test, increases in the maximum ALS plasma concentration (p = 0.0189) and the area under the plasma concentration–time curve from 0 to infinity (p = 0.0095). Conversely, a reduction was found in oral volume of distribution (p = 0.0009) and oral clearance (p = 0.0298). Blood urea nitrogen increased significantly after dosing in both groups (p < 0.0001 and p < 0.0001); however, no statistical difference was found between endpoint levels (p > 0.05) following one-way analysis of variance (ANOVA). Kidney injury molecule-1 increased following ALS dosing (p = 0.0486), while ALS-NP showed a decrease (p = 0.027) which was also significantly decreased compared to ALS-Final (p = 0.0008) when examined using two-way ANOVA. Urinary potassium excretion decreased significantly, as shown by two-way ANOVA, only in the ALS group (p = 0.0274) which was also significantly reduced compared to ALS-NP-Final (p = 0.016). Using the current formulation and at the dosage tested, ALS-NP showed a more favorable pharmacokinetic profile and positive kidney changes compared to ALS in regard to select outcomes. Thus, NP formulation may further improve ALS renoprotection in addition to increasing bioavailabilty.

Dissolved organic matter (DOM) plays a vital role in biogeochemical processes and can flux CO2 to the atmosphere when labile fractions are degraded, hence DOM degradation is increasingly studied. Some studies have suggested that fluorescence-derived substrate characteristics are useful metrics for estimating bioavailability (as prerequisite condition for biodegradability), however, recent findings on soil organic matter emphasize the importance of ecosystem scale factors such as physical separation of substrate from soil microbial communities and soil physiochemical cycles driving organic matter stability. I extend this principle to soil derived DOM and hypothesize that such environmental conditions, covariant with season, land use and landscape position, impact the composition of soil DOM and activity and abundance of the microbial community, which together govern DOM biodegradability. As a result, DOM bioavailability may not reliably be predicted using substrate characteristics alone. To test these hypotheses, I assessed aqueous soil extracts for water extractable organic carbon (WEOC) content, biodegradability, microbial biomass and fluorescence spectroscopy on water extractable organic matter (WEOM) across a range of environmental conditions in northern Vermont, USA. My results indicate that changes in environmental conditions affect composition, quantity, and biodegradability of DOM. WEOC concentrations were highest in the fall and lowest in the summer, while no significant differences were found between land covers or landscape position, however, DOM biodegradability was significantly higher in the agricultural (AG) site across seasons. Despite a shift in utilized substrate from less aromatic DOM in summer to more aromatic DOM in winter, biodegradability was similar for all seasons. The only exception were cold temperature incubations where microbial activity was depressed, and processing was halted. These results indicate that bioavailability cannot be reliably predicted based on fluorescence-based metric alone, rather, my core findings illustrate a complex picture of how environmental conditions, landscape characteristics, and substrate composition interact to drive the biodegradability of labile carbon pools in the soil environment. This thesis includes i) a background and comprehensive review of literature to inform the reader of any relevant topics, ii) a paper submitted for publication to Biogeochemistry (Chapter 2), and iii) supplemental information containing figures and tables pertinent to the paper.

The effect of iron absorption on subsequent hemoglobin regeneration was investigated by repletion and subsequent analysis of an anemic weanling rat model. Primary assay procedure in these studies was the Regeneration Efficiency method. As repletion or curative assay techniques form the central body of investigative research in iron utilization, the first subject of inquiry was the effect of repletion interval and degree of depletion upon the animals as monitored by physical indicators such as blood volume, growth and hematinic response. Percent blood volume is a particularly useful figure in the calculation of the amount of hemoglobin iron but was found to be relatively constant in a preliminary experiment. Accordingly, the effects of growth and anemia on hemoglobin response and blood volume were examined in 122 weanling male rats which had been depleted for seven days by low iron diet and phlebotomy, then repleted by feeding iron rich diet (47.1 ppm) for zero, five, 10 and 15 days. Percent blood volume proved to be rather constant at 7.5 in these pre-pubescent animals. The most severely depleted animals appeared to regenerate hemoglobin initially rather than replenish tissue iron. Regeneration Efficiency and AOAC assay methods were compared in a study involving the relative potency of two iron salts, ferrous sulfate and ferric orthophosphate, and three commercial cereal sources containing electrolytically reduced, hydrogen reduced or ferric orthophosphate supplements. These two assays utilized 202 albino male weanling rats. The superior availability of ferrous sulfate was evident in both regeneration efficiency and AOAC assays, as was the relative similarity of the dietary iron sources, yet the commercial sources in general provided more available iron in comparison to the reference ferrous sulfate than expected, possibly due to the influence of dietary components and processing variables. Bioavailability based on dietary iron concentration response correlated highly (r = 0.94) between animal groups analyzed using the Regeneration Efficiency and AOAC methods. Stress may have been a factor in animals assayed by the AOAC technique which used a 28 day depletion interval followed by a two week repletion. The Regeneration Efficiency method, which utilizes shorter depletion and repletion intervals also offered sufficient amounts of dietary iron for normal growth. The calculation of iron bioavailability also is dependent on such pertinent variables as dietary intake amount, body weight gain and percent blood volume.

<p>Mercury (Hg) pollution is a global environmental problem. Numerous Hg-contaminated sites exist in the world and new techniques for remediation are urgently needed. Phytoremediation, use of plants to remove pollutants from the environment or to render them harmless, is considered as an environment-friendly method to remediate contaminated soil <i>in-situ</i> and has been applied for some other heavy metals. Whether this approach is suitable for remediation of Hg-contaminated soil is, however, an open question. The aim of this thesis was to study the fate of Hg in terrestrial plants (particularly the high biomass producing willow, <i>Salix spp</i>.) and thus to clarify the potential use of plants to remediate Hg-contaminated soils.</p><p>Plants used for phytoremediation of Hg must tolerate Hg. A large variation (up to 30-fold difference) was detected among the six investigated clones of willow in their sensitivity to Hg as reflected in their empirical toxicity threshold (TT_95b), the maximum unit toxicity (UTmax) and EC50 levels. This gives us a possibility to select Hg-tolerant willow clones to successfully grow in Hgcontaminated soils for phytoremediation.</p><p>Release of Hg into air by plants is a concern when using phytoremediation in practice. No evidence was found in this study that Hg was released to the air via shoots of willow, garden pea (<i>Pisum sativum</i> L. cv Faenomen), spring wheat (<i>Triticum aestivum</i> L. cv Dragon), sugar beet (<i>Beta vulgaris </i>L. cv Monohill), oil-seed rape (<i>Brassica napus</i> L. cv Paroll) and white clover (<i>Trifolium repens</i> L.). Thus, we conclude that the Hg burden to the atmosphere via phytoremediation is not increased.</p><p>Phytoremediation processes are based on the ability of plant roots to accumulate Hg and to translocate it to the shoots. Willow roots were shown to be able to efficiently accumulate Hg in hydroponics, however, no variation in the ability to accumulate was found among the eight willow clones using CVAAS to analyze Hg content in plants. The majority of the Hg accumulated remained in the roots and only 0.5-0.6% of the Hg accumulation was translocated to the shoots. Similar results were found for the five common cultivated plant species mentioned above. Moreover, the accumulation of Hg in willow was higher when being cultivated in methyl-Hg solution than in inorganic Hg solution, whereas the translocation of Hg to the shoots did not differ.</p><p>The low bioavailability of Hg in contaminated soil is a restricting factor for the phytoextraction of Hg. A selected tolerant willow clone was used to study whether iodide addition could increase the plant-accumulation of Hg from contaminated soil. Both pot tests and field trials were carried out. Potassium iodide (KI) addition was found to mobilize Hg in contaminated soil and thus increase the bioavailability of Hg in soils. Addition of KI (0.2–1 mM) increased the Hg concentrations up to about 5, 3 and 8 times in the leaves, branches and roots, respectively. However, too high concentrations of KI were toxic to plants. As the majority of the Hg accumulated in the roots, it might be unrealistic to use willow for phytoextraction of Hg in practice, even though iodide could enhance the phytoextraction efficiency.</p><p>In order to study the effect of willow on various soil fractions of Hg-contaminated soil, a 5-step sequential soil extraction method was used. Both the largest Hg-contaminated fractions, i.e. the Hg bound to residual organic matter (53%) and sulphides (43%), and the residual fraction (2.5%), were found to remain stable during cultivations of willow. The exchangeable Hg (0.1%) and the Hg bound to humic and fulvic acids (1.1%) decreased in the rhizospheric soil, whereas the plant accumulation of Hg increased with the cultivation time. The sum of the decrease of the two Hg fractions in soils was approximately equal to the amount of the Hg accumulated in plants. Consequently, plants may be suitable for phytostabilization of aged Hg-contaminated soil, in which root systems trap the bioavailable Hg and reduce the leakage of Hg from contaminated soils.</p>

Mercury (Hg) pollution is a global environmental problem. Numerous Hg-contaminated sites exist in the world and new techniques for remediation are urgently needed. Phytoremediation, use of plants to remove pollutants from the environment or to render them harmless, is considered as an environment-friendly method to remediate contaminated soil in-situ and has been applied for some other heavy metals. Whether this approach is suitable for remediation of Hg-contaminated soil is, however, an open question. The aim of this thesis was to study the fate of Hg in terrestrial plants (particularly the high biomass producing willow, Salix spp.) and thus to clarify the potential use of plants to remediate Hg-contaminated soils. Plants used for phytoremediation of Hg must tolerate Hg. A large variation (up to 30-fold difference) was detected among the six investigated clones of willow in their sensitivity to Hg as reflected in their empirical toxicity threshold (TT95b), the maximum unit toxicity (UTmax) and EC50 levels. This gives us a possibility to select Hg-tolerant willow clones to successfully grow in Hgcontaminated soils for phytoremediation. Release of Hg into air by plants is a concern when using phytoremediation in practice. No evidence was found in this study that Hg was released to the air via shoots of willow, garden pea (Pisum sativum L. cv Faenomen), spring wheat (Triticum aestivum L. cv Dragon), sugar beet (Beta vulgaris L. cv Monohill), oil-seed rape (Brassica napus L. cv Paroll) and white clover (Trifolium repens L.). Thus, we conclude that the Hg burden to the atmosphere via phytoremediation is not increased. Phytoremediation processes are based on the ability of plant roots to accumulate Hg and to translocate it to the shoots. Willow roots were shown to be able to efficiently accumulate Hg in hydroponics, however, no variation in the ability to accumulate was found among the eight willow clones using CVAAS to analyze Hg content in plants. The majority of the Hg accumulated remained in the roots and only 0.5-0.6% of the Hg accumulation was translocated to the shoots. Similar results were found for the five common cultivated plant species mentioned above. Moreover, the accumulation of Hg in willow was higher when being cultivated in methyl-Hg solution than in inorganic Hg solution, whereas the translocation of Hg to the shoots did not differ. The low bioavailability of Hg in contaminated soil is a restricting factor for the phytoextraction of Hg. A selected tolerant willow clone was used to study whether iodide addition could increase the plant-accumulation of Hg from contaminated soil. Both pot tests and field trials were carried out. Potassium iodide (KI) addition was found to mobilize Hg in contaminated soil and thus increase the bioavailability of Hg in soils. Addition of KI (0.2–1 mM) increased the Hg concentrations up to about 5, 3 and 8 times in the leaves, branches and roots, respectively. However, too high concentrations of KI were toxic to plants. As the majority of the Hg accumulated in the roots, it might be unrealistic to use willow for phytoextraction of Hg in practice, even though iodide could enhance the phytoextraction efficiency. In order to study the effect of willow on various soil fractions of Hg-contaminated soil, a 5-step sequential soil extraction method was used. Both the largest Hg-contaminated fractions, i.e. the Hg bound to residual organic matter (53%) and sulphides (43%), and the residual fraction (2.5%), were found to remain stable during cultivations of willow. The exchangeable Hg (0.1%) and the Hg bound to humic and fulvic acids (1.1%) decreased in the rhizospheric soil, whereas the plant accumulation of Hg increased with the cultivation time. The sum of the decrease of the two Hg fractions in soils was approximately equal to the amount of the Hg accumulated in plants. Consequently, plants may be suitable for phytostabilization of aged Hg-contaminated soil, in which root systems trap the bioavailable Hg and reduce the leakage of Hg from contaminated soils.

Electrodialysis (ED) experiments were conducted on reverse osmosis (RO)-concentrated solutions of NOM from six rivers. The ED processes successfully recovered 88 11% of TOC, and removed 83% 19% of SO42- and 67% 18% of H4SiO4. More importantly, the molar ratios of SO42- /TOC and H4SiO4 /TOC were reduced to a mean value of 0.0046 and 0.032, respectively, surpassing the goal for removal of SO42- (0.008) and almost achieving the goal for removal of H4SiO4 (0.021). The ED process can lower the SO42- /TOC ratio in samples whose initial SO42- /TOC ratios are already far below the limit of 0.008 used in this study. The coupled RO/ED process that has been described here offers a fast, simple, chemically mild (relative to other methods), and reproducible method of isolation of large quantities of relatively unfractionated, low-ash NOM from freshwaters. RO/ED was also successfully used for isolating and concentrating marine dissolved organic matter (DOM). The effort successfully recovered a median of 72% of the TOC from 200 L samples within six to nine hours of processing through a combination of ED and RO, greatly exceeding the current norm of 30%. The relatively high recovery of DOM implies that classes of DOM previously missing are included in these samples and should yield new insight into the chemistry of marine DOM. Freshwater samples processed by electrodialysis were analyzed for elemental composition and by capillary zone electrophoresis (CZE), 1H and 13C nuclear magnetic resonance spectroscopy (NMR), and electro-spray ionization mass spectrometry (ESI-MS). Bulk elemental composition, 1H- and 13C-NMR, and ESI-MS data provide evidence linking bioavailabilty to the bulk chemistry of NOM: the H/C and N/C molar ratios are positively and strongly correlated with bioavailability, as hypothesized. Using an independent dataset (STORET) of water quality parameters, calculated BOD/TOC ratios were found to be moderately correlated with measured bioavailabilities and can be used as a surrogate for bioavailability of geochemically diverse riverine DOM.

Diss. (sammanfattning) Uppsala : Sveriges lantbruksuniv., 2009. / Härtill 4 uppsatser.

Not available / text

DL-Methionine (Met) has been conventionally used in swine diets with assumption of similar bioefficacy with L-Met. However, because L-Met is the form that is utilized by animals for protein synthesis, L-Met could, theoretically, be more available. Four experiments were conducted to evaluate L-Met bioavailability in nursery pigs with 21-day growth trials. A total of 105,105,112 and 84 crossbred pigs were used in Exp. 1, 2, 3 and 4, respectively. Each experiment had a low Met basal diet and 3 levels of the Met sources (DL-Met and L-Met). In addition to the basal diet, supplementation levels were 0.053%, 0.107% and 0.160% in Exp. 1, 0.040%, 0.080% and 0.120% in Exp. 2, 0.033%, 0.067% and 0.100% in Exp.3, 0.040%, 0.080% and 0.120% in Exp. 4. Body weight (BW), average daily gain (ADG), average daily feed intake (ADFI), gain: feed (G:F) were measured and plasma urea nitrogen (PUN) was analyzed in blood samples weekly. In Exp. 3 and 4, preference studies were conducted with the basal diet and the second highest level of each Met source. When additional DL-Met or L-Met were supplemented to the basal diet, BW, ADG, ADFI, and G:F ratio increased (P < 0.05). In the comparison between the DL-Met and L-Met diets in Exp. 1, pigs in the L-Met group had greater ADG and G:F ratios in the d 0-7 (P < 0.05) period than those in the DL-Met group. However, there were no differences for the overall experimental period. In Exp. 2, pigs in the DL-Met group had greater BW (P < 0.05), ADG (P < 0.05) and ADFI (P < 0.05) than those in the L-Met group for the overall period whereas no differences were observed in G:F ratios and PUN concentrations. In Exp. 3 and 4, there were no differences in BW, ADG, ADFI, G:F ratios or PUN concentrations between L-Met and DL-Met groups for the overall period. There was no preference exhibited for either the DL-Met or L-Met diet. In the results of relative bioavailability of L-Met to DL-Met, the values was 111.1% for d 0-14 based on the estimation by ADG in Exp. 1; L-Met bioavailability was lower than DL-Met based on all response measures in Exp. 2. However, in Exp. 3, relative bioavailability of L-Met to DL-Met was 100.4, 147.3, and 104.1% for d 0-14 ADG, G:F ratio and PUN concentrations. In Exp 4, the relative bioavailability of L-Met was 92.9, 139.4 and 70.4% for d 0-14 ADG, G:F ratio and PUN concentrations. In conclusion, using L-Met in the nursery diet demonstrated no consistent beneficial effect on ADG, G:F ratio or relative bioavailability compared to conventional DL-Met.

d-Limonene is a monoterpene found in high concentration in citrus peel oil. Evidence from animal models and cell culture indicate that it has strong anti-cancer effects, particularly in mammary cancer models. Chapter 1; "D-LIMONENE: A BIOACTIVE FOOD COMPONENT FROM CITRUS AND EVIDENCE FOR A POTENTIAL ROLE IN BREAST CANCER PREVENTION AND TREATMENT" is a review paper accepted to Oncology Reviews. This review describes the evidence for <italic>d<italic>-limonene's anti-cancer mechanisms, bioavailability and safety, focusing on relevance to breast cancer prevention. Chapter 2; "ADIPOSE TISSUE ACCUMULATION OF D-LIMONENE WITH THE CONSUMPTION OF A LEMONADE PREPARATION RICH IN D-LIMONENE CONTENT" is published in Nutrition and Cancer journal and describes a phase I clinical trial in which participants consumed 40 oz of high-limonene lemonade daily. This study demonstrated that after 4 weeks of oral consumption of high-limonene lemonade, d-limonene deposits in high levels in adipose tissue. Chapter 3; "A CLINICAL BIOMARKER STUDY OF TOPICALLY APPLIED D-LIMONENE FOR BREAST CANCER PREVENTION" was submitted to Nutrition and Cancer journal. In this phase 0 clinical study, four weeks of a 10% d-limonene formulation resulted in minimal change in NAF and plasma biomarkers or d-limonene levels. Biomarkers in NAF and plasma, however, were significantly differently correlated with BMI and menopausal status, perhaps suggesting effect modifications. Chapter 4: "MOUSE MAMMARY TISSUE DISTRIBUTION OF D-LIMONENE AND PERILLIC ACID FOLLOWING ORAL AND TOPICAL D-LIMONENE ADMINISTRATION," was a study comparing d-limonene and perillic acid disposition after administration of 10% and 20% d-limonene in coconut oil in topical and oral forms to SKH-1 mice. This study demonstrated that d-limonene deposits in high levels in mouse mammary tissue after both oral and topical administration short-term, but is largely cleared after 24 hours in this model. Perillic acid deposits in high levels in adipose after oral administration, and these high concentrations remained after 24 hours. Chapter 5: "IMPLICATIONS AND FUTURE DIRECTIONS" provides a summary of the key findings from these three projects and proposals for future research. The appendices provide results from smaller d-limonene projects, as well as extensions of the body of the dissertation work.