京都大学 / 新制・論文博士 / 博士(農学) / 乙第13413号 / 論農博第2896号 / 新制||農||1085(附属図書館) / 学位論文||R3||N5323(農学部図書室) / 京都大学大学院農学研究科農学専攻 / (主査)教授 松村 康生, 教授 白岩 立彦, 教授 丸山 伸之 / 学位規則第4条第2項該当 / Doctor of Agricultural Science / Kyoto University / DGAM

Electrospinning of nanofibrous mats and scaffolds enables generation of scaffolding that is not only highly porous, but also has a structure that essentially mimics the natural basement membrane. As a result, the method has proliferated extensively, and is commonly used for diverse applications such as water filtration or tissue engineering, the latter of which may involve the use of natural or synthetic materials. Common laboratory scale electrospinning setups can be built inexpensively with merely a syringe pump, a high voltage supply, and an aluminum foil target. These systems, however, are limited to flat target surface geometries that span several centimeters. While a scaffold can be cut or folded to conform to a bone or other biological surface, spinning directly onto a surface with significant peaks and troughs results in poor fiber uniformity. Furthermore, if an alteration of fiber properties is preferred, the high voltage setup limits user access and customization of parameters during the spinning period. Finally, control of the electric field is compromised by the proximity of grounded electrical components. As its first aim, this project develops a robotic control system to enable custom coatings of arbitrary surfaces. By augmenting the traditional electrospinning system with a three-dimensional robotic control system, electric field focusing fibers, and additional aerodynamic forces terms ‘electroblowing’, the device can be produced across targets with strong topographic anisotropy. The second aim continues to enhance these attributes with biocompatible soy based scaffolds. Craniofacial implants are often complex in geometry, and conformal bandages are particularly hard to produce in these areas. Soy based scaffolds will be produced for 3D-printed replicas of these situations. Finally, the methods developed across this aim enables the development and use of a handheld electrospinning system that combines a coaxial high velocity air flow with the high voltage spinning element to reduce effects of operator error. The final goal of the thesis is to test whether fiber control successfully reduces effects of fiber anisotropy in vitro and to use the enhanced fiber control mechanisms to produce scaffolds with significant anisotropy, depositing aligned fibers at a target point to eventually enable generation of scaffolds with programmable variable spatial alignment similar to tendon. When completed, the systems described will enable custom production of coatings or scaffolds for functionality as scaffolding on medically relevant surfaces. Specifically, this means first, that scaffolds can be used with confidence to improve fixation even of non-cylindrical implants and enhance local tissue integration, and second, that implants can be customized with areas of ‘guidance’ fibers or local drug depots to either promote regeneration and population by surrounding tissue or mimic natural anisotropic cues necessary for mechanical or biological functionality. / Bioengineering

Dietary soy isoflavones have been shown to ameliorate insulin resistance and Type 2 diabetes. However, many in vitro studies used supra-physiological concentrations of individual isoflavones that make it difficult to interpret the results as potential mechanisms in vivo. Since the insulin-sensitizing effects of thiazolidinediones, anti-diabetic drugs, have been shown to be mediated through activation of peroxisome proliferators-activated receptor gamma (PPARγ), the key transcription factor for adipocyte differentiation, we examined the effects of the two main soy isoflavones genistein and daidzein either as individual compound or combined on adipocyte differentiation and PPARγ expression, as well as whether the Wnt/β-catenin signaling pathway is the underlying molecular mechanism. In 3T3-L1 cells, genistein and daidzein significantly enhanced adipocyte differentiation. Similarly the expression of PPARγ increased particularly at 20 µmol/L. The stimulatory effect is greater when the two isoflavones are combined, indicating a synergistic effect. Genistein and daidzein also increased the relative abundance of insulin-responsive glucose transporter 4 (GLUT4) mRNA with a greater effect when combined. Wnt10b expression was not affected by soy isoflavones treatments, while Wnt5b expression was only increased by the combination of genistein and daidzein. Our results suggest, that the combination of soy isoflavones has a greater effect in increasing the newly formation of adipocytes that are highly insulin-sensitive via an increase in PPARγ expression as well as increasing the expression of GLUT4. However, genistein and daidzein actions on Wnt signaling remain unclear. These data further support the epidemiological findings for the beneficial effect of soy consumption on insulin sensitivity.

Low moisture veneer and regions of sapwood within hickory engineered wood flooring bonded with soy-flour adhesive are thought to be factors leading to potential performance deficiencies. The goal of this research was to gain a broader understanding of bonding low moisture hickory veneer with soy-based adhesive. Soyad® is of particular interest due to its novel cross-linking chemistry. Impacts of moisture content and wood region (heartwood versus sapwood) were analyzed with dry and wet shear bond strength tests, measurement of percent wood failure, lathe check characterization, and adhesive bondline thickness and penetration depth measurement. Impact of wood region and type (hickory versus red oak) was assessed by comparing wood buffering capacity and delamination following three-cycle water soaking. Dry and wet shear strength values met expectations for engineered wood flooring yet percentage wood failure results were uniformly very low for all combinations of moisture levels and wood regions. In contrast, delamination following wet and dry cycling was minor and within minimum requirements for all specimens tested. The influence of moisture level, wood region and type were inconsistent; statistically significant relationships were not evident within the moisture range studied. However, different wood regions and types exhibited differing veneer buffering capacities that had potential to interfere with pH requirements of Soyad®. Additional study of buffering capacity and resin cure is recommended to determine the significance of the buffering capacity results found in this study. / Master of Science / Performance issues including debonding and delamination have been reported for hickory engineered wood flooring products constructed using a soy-flour based adhesive. Sapwood regions within the composite and extremely low moisture veneer were provided by industry as possible factors that resulted in performance deficiencies. The goal of this research project was to gain a broader understanding of bonding low moisture hickory veneer with Soyad® adhesive. Soy-flour adhesive systems offer many environmental, health, and durability advantages. Soyad® is of particular interest due to its use of natural, renewable soy flour, a novel cross-linking resin, and no added formaldehyde. Test specimens were prepared using heartwood of hickory and red oak and sapwood of hickory. Analytical tests included determination of certain chemical properties of the adhesive and wood veneer, measurement of strength properties of the adhesive bond, and assessment of delamination tendencies of bonded panels following water soaking. Results indicate that moisture levels and the different growth regions and wood type had an inconsistent impact on the bond strengths yet percent wood failure was uniformly low and considered unacceptable by industry. Although this research established a foundation of basic knowledge, more information about adhesive bonding of wood with the recently developed soy based adhesives is needed to optimize the systems and provide technological advancements that lead to more efficient and safe utilization of woody materials from the forest.

L to D isomerization of amino acids in a commercially prepared alkali-treated food-grade soy isolate was investigated. A soy flour product was also investigated as a control which had not been alkali treated. Methionine and valine collected from two soy flour and two soy isolate samples were coupled with L-leucine-N-carboxyanhydride. If both D- and L-forms were present this would produce diastereomeric dipeptide derivatives. The derivatives were separated by ion-exchange chromatography using a TSM Amino Acid Analyzer. Comparison of the sample methionine and valine dipeptide peaks to standard dipeptide peaks revealed the presence of L-valine, L-methionine and D-methionine in both soy flour and soy isolate samples. Methionine was substantially isomerized in both products. D-valine was not observed in either soy flour or soy isolate samples. Thus, isomerization of methionine appeared to occur at a faster rate than valine. Acid hydrolysis, toasting of soy meal, and alkaline treatment are all conditions which may be responsible for the isomerization of methionine. / Master of Science

Building waste and disposable packaging are a major component in today's landfills. Most of these are structural or thermally insulative polymer foams that do not degrade over a long period of time. Currently, there is a push to replace these foams with thermoplastic or biodegradable foams that can either be recycled or composted. We propose the use of compostable soy-based polyurethane foams (PU) with kenaf core modifiers that will offer the desired properties with the ability to choose responsible end-of-life decisions. The effect of fillers is a critical parameter in investigating the thermal and mechanical properties along with its effect on biodegradability. In this work, foams with 5%, 10%, and 15% kenaf core content were created. Two manufacturing approaches were used: the free foaming used by spray techniques and the constrained expansion complementary to a mold cavity. Structure-property relations were examined using differential scanning calorimetry (DSC), dynamic mechanical analysis (DMA), thermal conductivity, compression values, scanning electron microscopy (SEM), x-ray micro-computed tomography (micro-CT), and automated multiunit composting system (AMCS). The results show that mechanical properties are reduced with the introduction of kenaf core reinforcement while thermal conductivity and biodegradability display a noticeable improvement. This shows that in application properties can be improved while establishing a responsible end-of-life choice.

Lors du développement et de l’introduction d’un nouvel aliment, la compréhension des croyances, attitudes et préférences sensorielles des consommateurs est très importante. Cette thèse présente une série d'études cross-culturelles, France vs. Vietnam, sur la représentation des consommateurs vis-à-vis de produits à base soja et sur la perception et la préférence des yaourts à base de soja, un produit dérivé du soja récemment développé pour le marché français et n'existant pas sur le marché vietnamien. Malgré une image favorable du soja sur le plan nutritionnel et sur la santé, les participants français ont rapporté ne pas consommer ce type de produit, la barrière à sa consommation étant son goût de soja. Ce résultat a été confirmé par un test de consommateurs au cours duquel les participants français ont donné une très faible évaluation hédonique pour les yaourts de soja. Les consommateurs vietnamiens, consommateurs traditionnels du soja, n'ont également pas apprécié ces produits. Une série de yaourts mixtes dans lesquels le lait de vache a été partiellement remplacé par celui de soja a été fabriquée. Les tests consommateurs ont montré que pour les deux pays, le yaourt fabriqué avec une proportion de lait de soja inférieure à 50% a reçu une note moyenne supérieure à 5 sur une échelle de 1 à 9. Le rôle de la familiarité sur les préférences alimentaires ainsi que l'impact de l'exposition culturelle sur les changements des croyances et attitudes à l'âge adulte ont été confirmés. La présente étude contribue à la caractérisation des deux cultures sur leur acceptabilité d'un nouvel aliment / Understanding consumers’ beliefs, attitudes and sensory preference is the key to a successful development and introduction of a new food. This thesis carried out a series of cross-cultural studies, France vs. Vietnam, on consumers’ representation of soy foods and on sensory perception and preference of soy yogurts, a soy derived product recently developed for French market which does not exist on the Vietnamese market. Despite a favourable image of soy in terms of nutrition and health benefices, French participants reported that the barrier to its consumption was the taste of soy. This finding was confirmed by a consumer test in which French participants gave very low hedonic evaluations to soy yogurts. Interestingly, Vietnamese consumers, soy traditional consumers, did not appreciate these products as well. A series of mix yogurts in which cow milk was partially replaced by soy milk was formulated. Consumer tests showed that for both countries, a mix yogurt with proportion of soy milk lower than 50% received an average hedonic score higher than 5 on a scale going from 1 to 9. The role of the familiarity on food preference and the impact of cultural exposure on changing in beliefs and attitudes’ change at adult age were confirmed. The present study contributes to the characterisation of the two cultures on their acceptability of a new food

Background: Bacteriophage Φ6 is a lipid-enveloped dsRNA bacteriophage. The limitations in our knowledge of how this bacteriophage occurs in the environment are limited by non-selective isolation techniques. Research on finding phages in the environment in the past has employed the Double Agar Layer (DAL) plaque assay using Tryptic Soy Agar (TSA), a non-selective media. The bacterial host for bacteriophage Φ6 is Pseudomonas syringae. In this study, we tested Pseudomonas Agar, a selective media that suppresses the growth of bacteria except Pseudomonas species, in the standard double agar layer plaque assay for Φ6. Methods: DAL plaque assays were performed to determine the sensitivity of both Tryptic Soy Agar (TSA) and Pseudomonas Agar (PA) for determining the titer of pure bacteriophage Φ6 stocks. We used Pseudomonas syringae (HB10Y) as the host, and the plaque formation on both agars was compared. Following the evaluation of PA with pure Φ6 stocks, PA effectiveness for Φ6 isolation from environmental samples was tested in spiked waters obtained from irrigation ponds at an agricultural farm. Results: Comparison of TSA and PA using pure Φ6 cultured in the laboratory and spiked environmental samples showed that PA agar can detect bacteriophage Φ6 as well as the standard DAL assay using TSA. On PA, formation of clear visible plaques comparable to the plaques formed using TSA was observed. Conclusions: Pseudomonas Agar can be used for the isolation of bacteriophage Φ6 in environmental samples. This may enhance the detection of these phages in the environment.

The objective of this study was to determine if acoustic emissions (AE) generated by three strains of Escherichia Coli (5024-parent strain, 8279-mutant strain and 8279-random/unrelated strain) could be used to differentiate each strain during their growth cycle. An acoustic sensor with an operating range of 35 kHz-100 kHz was inserted into the growth vessel and attached to a selected channel to capture AE data. The growth vessel was loaded with 60 ml of tryptic soy broth (TSB) (0.25% fructose) media with alginate (1.1%) or without alginate and inoculated with 1% (108 CFU/ml) of an E. coli strain. The growth vessel was placed in a monitoring chamber and incubated at 32°C for 8-9 h. The AE’s generated by each strain were collected throughout the growth cycle. All strains grown in media with and without alginate generated AE’s within 5 min post inoculation. Strains grown in media without alginate generated stronger (P < 0.0001) absolute energy (ABSE) and higher peak frequencies (PFRQ’s), than in media with alginate. The AE’s generated by strains 5024 and 8237 were stronger and easily distinguished from those generated by strain 8279. Strain 8237 generated 12% stronger ABSE from the 3rd to 8th h and 51% stronger PFRQ intensities than strain 5024 during 0-8 h. However, strain 5024 generated 15% stronger ABSE and 31% higher PFRQ’s during the final hour of growth. Strain 5024 generated the highest PFRQ’s from 5-50 kHz, while strain 8237 generated higher frequencies from 100-500 kHz. Fourteen distinguishable differences (P< 0.05) in generated PFRQ’s, between strains 5024 and 8237, were also observed in every 5 kHz increments from 100-500 kHz. Of these differences, strain 8237 generated higher frequencies within eight of the kHz ranges, while strain 5024 generated higher frequencies within six other kHz ranges. These data suggests that all bacteria may generate different AE’s, thus producing a unique “fingerprint” of sound that will allow for its identification.

Master of Science / Department of Grain Science and Industry / Sajid Alavi / Extrusion processing and frying are the two most commonly used methods to produce savory snacks. These snacks are mostly starch based and also contain high amounts of fat. Snacking percentage has increased drastically over the past few decades causing many health problems like obesity, hypertension and cardiovascular diseases. The overall objective of this study was to develop novel soy based savory snack that are high in protein and has less fat using both extrusion processing and frying. In the first part of this study both defatted soy (25-75%), wheat flour, monoglycerides (0.375 & 0.75%) and sodium bicarbonate (0 and 0.5%) were used to produce dense extrudates, which mimic the shape of lentils using pilot scale twin screw extruder. The extrudates were soaked in water and fried in corn oil to produced savory snacks. Soy influenced the water absorption during soaking and oil uptake during frying. The water uptake and % fat decreased with increase in soy and the amount of water uptake influenced the % fat absorption in the product. No significant differences were observed in overall acceptability and to summarize the increased protein and dense structure of the products challenge the texture of the products. In the second part of this research, dynamics of water and oil uptake were studied to see the role of texture modifiers like soy protein isolate (4, 8, & 12%), calcium bicarbonate (0.2, 0.4 & 0.6%) and pregelatinized wheat starch (4, 8, & 12%) on texture, water and oil uptake. Descriptive sensory analysis was conducted to study the sensory attributes of the products. Water holding capacity is influenced by the level of % starch addition and the degree of starch degradation during processing. Oil uptakes correlates to that of WHC and is also affected by the degree of gelatinization and crust formation. Degree of starch gelatinization, oil uptake and oil and starch matrix interactions had an impact on hardness. Among SPI, CaCO3 and PGWS, PGWS lowered the product hardness followed by SPI and CaCO3. Descriptive sensory results are similar to experimental results with PGWS samples having lower initial or substantial hardness and more oxidized or heated oil aroma and flavor.