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Optimization and single-laboratory validation of a method for the determination of flavonolignans in milk thistle seeds by high-performance liquid chromatography with ultraviolet detection
Seeds of milk thistle, Silybum marianum (L.) Gaertn., are used for treatment and prevention of liver disorders and were identified as a high priority ingredient requiring a validated analytical method. An AOAC International expert panel reviewed existing methods and made recommendations concerning method optimization prior to validation. A series of extraction and separation studies were undertaken on the selected method for determining flavonolignans from milk thistle seeds and finished products to address the review panel recommendations. Once optimized, a single-laboratory validation study was conducted. The method was assessed for repeatability, accuracy, selectivity, LOD, LOQ, analyte stability, and linearity. Flavonolignan content ranged from 1.40 to 52.86 % in raw materials and dry finished products and ranged from 36.16 to 1570.7 μg/mL in liquid tinctures. Repeatability for the individual flavonolignans in raw materials and finished products ranged from 1.03 to 9.88 % RSD, with HorRat values between 0.21 and 1.55. Calibration curves for all flavonolignan concentrations had correlation coefficients of >99.8 %. The LODs for the flavonolignans ranged from 0.20 to 0.48 μg/mL at 288 nm. Based on the results of this single-laboratory validation, this method is suitable for the quantitation of the six major flavonolignans in milk thistle raw materials and finished products, as well as multicomponent products containing dandelion, schizandra berry, and artichoke extracts. It is recommended that this method be adopted as First Action Official Method status by AOAC International., Peer-reviewed article, Published. Received 1 June 2015; Revised 14 July 2015; Accepted 16 July 2015; Published online 31 July 2015.
Osmoregulation in the halophilic bacterium halomonas elongata
Halophilic bacteria use a variety of osmoregulatory methods, such as the accumulation of one or more compatible solutes. The wide diversity of compounds that can act as compatible solute complicates the task of understanding the different strategies that halophilic bacteria use to cope with salt. This is specially challenging when attempting to go beyond the pathway that produces a certain compatible solute towards an understanding of how the metabolic network as a whole addresses the problem. Metabolic reconstruction based on genomic data together with Flux Balance Analysis (FBA) is a promising tool to gain insight into this problem. However, as more of these reconstructions become available, it becomes clear that processes predicted by genome annotation may not reflect the processes that are active in vivo. As a case in point, E. coli is unable to grow aerobically on citrate in spite of having all the necessary genes to do it. It has also been shown that the realization of this genetic potential into an actual capability to metabolize citrate is an extremely unlikely event under normal evolutionary conditions. Moreover, many marine bacteria seem to have the same pathways to metabolize glucose but each species uses a different one. In this work, a metabolic network inferred from genomic annotation of the halophilic bacterium Halomonas elongata and proteomic profiling experiments are used as a starting point to motivate targeted experiments in order to find out some of the defining features of the osmoregulatory strategies of this bacterium. This new information is then used to refine the network in order to describe the actual capabilities of H. elongata, rather than its genetic potential., Peer-reviewed article, Published. Received: September 20, 2016; Accepted: November 17, 2016; Published: January 12, 2017.
The path of the smart grid
Exciting yet challenging times lie ahead. The electrical power industry is undergoing rapid change. The rising cost of energy, the mass electrification of everyday life, and climate change are the major drivers that will determine the speed at which such transformations will occur. Regardless of how quickly various utilities embrace smart grid concepts, technologies, and systems, they all agree onthe inevitability of this massive transformation. It is a move that will not only affect their business processes but also their organization and technologies., Final article published
Pattern selection in plants
Background and Aims A study is made by computation of the interplay between the pattern formation of growth catalysts on a plant surface and the expansion of the surface to generate organismal shape. Consideration is made of the localization of morphogenetically active regions, and the occurrence within them of symmetry-breaking processes such as branching from an initially dome-shaped tip or meristem. Representation of a changing and growing three-dimensional shape is necessary, as two-dimensional work cannot distinguish, for example, formation of an annulus from dichotomous branching. Methods For the formation of patterns of chemical concentrations, the Brusselator reaction-diffusion model is used, applied on a hemispherical shell and generating patterns that initiate as surface spherical harmonics. The initial shape is hemispherical, represented as a mesh of triangles. These are combined into finite elements, each made up of all the triangles surrounding each node. Chemical pattern is converted into shape change by moving nodes outwards according to the concentration of growth catalyst at each, to relieve misfits caused by area increase of the finite element. New triangles are added to restore the refinement of the mesh in rapidly growing regions. Key Results The postulated mechanism successfully generates: tip growth (or stalk extension by an apical meristem) to ten times original hemisphere height; tip flattening and resumption of apical advance; and dichotomous branching and higher-order branching to make whorled structures. Control of the branching plane in successive dichotomous branchings is tackled with partial success and clarification of the issues. Conclusions The representation of a growing plant surface in computations by an expanding mesh that has no artefacts constraining changes of shape and symmetry has been achieved. It is shown that one type of pattern-forming mechanism, Turing-type reaction-diffusion, acting within a surface to pattern a growth catalyst, can generate some of the most important types of morphogenesis in plant development., Peer-reviewed article, Published. Received: 26 July 2007; Returned for revision: 5 October 2007; Accepted: 15 October 2007; Published electronically: 28 November 2007.
Performance of sprayed fiber reinforced polymer strengthened timber beams
A study was carried out to investigate the use of Sprayed Fiber Reinforced Polymer (SFRP) for retrofit of timber beams. A total of 10-full scale specimens were tested. Two different timber preservatives and two different bonding agents were investigated. Strengthening was characterized using load deflection diagrams. Results indicate that it is possible to enhance load-carrying capacity and energy absorption characteristics using the technique of SFRP. Of the two types of preservatives investigated, the technique appears to be more effective for the case of creosote-treated specimens, where up to a 51% improvement in load-carrying capacity and a 460% increase in the energy absorption capacity were noted. Effectiveness of the bonding agent used was dependent on the type of preservative the specimen had been treated with., Peer-reviewed article, Published. Received 26 July 2010; Revised 8 October 2010; Accepted 12 October 2010.
Phase change material's (PCM) impacts on the energy performance and thermal comfort of buildings in a mild climate
The current residential buildings are of light weight construction. As such, they tend to frequent indoor air temperatures fluctuations and have been proven detrimental for thermal comfort and mechanical system energy consumption. This is reflected in the energy consumption statistics for residential buildings. More than 62% of the building energy use is towards maintaining comfortable indoor conditions. Phase change materials (PCM); a latent heat thermal storage material, have the potential to increase the thermal mass of these buildings without drastically affecting the current construction techniques. In this paper, the potential of phase change materials is investigated through numerical and experimental studies. The field experimental study is conducted using twin side-by-side buildings exposed to the same interior and exterior boundary conditions, and EnergyPlus, after being benchmarked with the experimental results, is used for the numerical study. The numerical study is carried out for an existing residential apartment unit with particular emphasis on the effects of different design parameters such as orientation and window to wall ratio. Preliminary analyses of experimental data show that phase change materials are effective in stabilizing the indoor air by reversing the heat flow direction. In fact, the indoor air and wall temperature fluctuations are reduced by 1.4 °C and 2.7 °C respectively. Following, benchmarking of the numerical simulation shows confidence levels in predicting the interior conditions since discrepancies between experimental data and numerical data are within tolerance limits of the measuring device. Further, from the analysis of the numerical data, phase change material is effective in moderating the operative temperature but does not translate to significant thermal comfort improvement when evaluated over a night time occupancy regime in the summer. On the contrary, PCM is effective in lowering the heating energy demand by up to 57% during the winter condition., Peer reviewed article, Published. Received 1 October 2015, Revised 22 January 2016, Accepted 23 January 2016, Available online 29 January 2016.
Planning for climate action in British Columbia, Canada
Significant greenhouse gas (GHG) reductions from all sectors of human enterprise are necessary to avoid further effects and reduce the current effects of climate change. Agriculture and the global food system are estimated to contribute to one-third of all anthropogenic GHGs. In British Columbia, Canada, mandated GHG reduction targets and voluntary climate action programs are challenging local governments to include emission reduction targets, policies, and actions within official planning documents. At this early stage of GHG reductions, local government attention does not yet include agriculture but is directed toward the transportation, buildings, and waste management sectors. Given agriculture's contribution to GHG emissions and local government's engagement with GHG mitigation and food system planning, it seems reasonable to anticipate that over time, local governments should and will engage increasingly in reducing GHGs from agriculture. With the goal of advancing agriculture GHG mitigation by local governments, this paper reviews the jurisdictional powers governing agriculture and climate change within British Columbia. It examines how local governments can support mitigation within the sector through their roles in planning, policy, programming, and public engagement, and identifies potential research agenda items., Peer-reviewed article, Published. Submitted 18 April 2011 ; Revised 4 July 2011 and 1 August 2011 ; Accepted 2 September 2011 ; Published online 20 March 2012.
Precision of non-invasive temperature measurement by diffuse reflectance spectroscopy
Diffuse reflectance spectroscopy can be used as a noninvasive probe for measurement of temperature in real time. We have measured the precision of this technique to determine the temperature of Si and GaAs substrates during semiconductor processing. Our results show that the standard deviation of the noninvasive optical technique is less than 1.5 °C for GaAs and less than 2.0 °C for Si over the temperature range 25 °C≪T≪600 °C. This standard deviation compares favorably to that for a type‐K thermocouple used in the same measurements: s.d.≪1.5 °C. These results support the notion that noninvasive optical temperature measurement can be used in semiconductor processing with a precision approaching that of a thermocouple., Peer-reviewed article, Published.
Proteomic analysis of the effects of aged garlic extract and its FruArg component on lipopolysaccharide-induced neuroinflammatory response in microglial cells
Aged garlic extract (AGE) is widely used as a dietary supplement, and is claimed to promote human health through anti-oxidant/anti-inflammatory activities with hypolipidemic, antiplatelet and neuroprotective effects. Prior studies of AGE have mainly focused on its organosulfur compounds, with little attention paid to its carbohydrate derivatives, such as N-α-(1-deoxy-D-fructos-1-yl)-L-arginine (FruArg). The goal of this study is to investigate actions of AGE and FruArg on antioxidative and neuroinflammatory responses in lipopolysaccharide (LPS)-activated murine BV-2 microglial cells using a proteomic approach. Our data show that both AGE and FruArg can significantly inhibit LPS-induced nitric oxide (NO) production in BV-2 cells. Quantitative proteomic analysis by combining two dimensional differential in-gel electrophoresis (2D-DIGE) with mass spectrometry revealed that expressions of 26 proteins were significantly altered upon LPS exposure, while levels of 20 and 21 proteins exhibited significant changes in response to AGE and FruArg treatments, respectively, in LPS-stimulated BV-2 cells. Notably, approximate 78% of the proteins responding to AGE and FruArg treatments are in common, suggesting that FruArg is a major active component of AGE. MULTICOM-PDCN and Ingenuity Pathway Analyses indicate that the proteins differentially affected by treatment with AGE and FruArg are involved in inflammatory responses and the Nrf2-mediated oxidative stress response. Collectively, these results suggest that AGE and FruArg attenuate neuroinflammatory responses and promote resilience in LPS-activated BV-2 cells by suppressing NO production and by regulating expression of multiple protein targets associated with oxidative stress., Peer-reviewed article, Published. Received: May 16, 2014; Accepted: October 24, 2014; Published: November 24, 2014.
Protracted myelin clearance hinders central primary afferent regeneration following dorsal rhizotomy and delayed neurotrophin-3 treatment
Regeneration within or into the CNS is thwarted by glial inhibition at the site of a spinal cord injury and at the dorsal root entry zone (DREZ), respectively. At the DREZ, injured axons and their distal targets are separated by degenerating myelin and an astrocytic glia limitans. The different glial barriers to regeneration following dorsal rhizotomy are temporally and spatially distinct. The more peripheral astrocytic barrier develops first, and is surmountable by neurotrophin-3 (NT-3) treatment; the more central myelin-derived barrier, which prevents dorsal horn re-innervation by NT-3-treated axons, becomes significant only after the onset of myelin degeneration. Here we test the hypothesis that in the presence of NT-3, axonal regeneration is hindered by myelin degeneration products. To do so, we used the Long Evans Shaker (LES) rat, in which oligodendrocytes do not make CNS myelin, but do produce myelin-derived inhibitory proteins. We show that delaying NT-3 treatment for 1 week in normal (LE) rats, while allowing axonal penetration of the glia limitans and growth within degenerating myelin, results in misdirected regeneration with axons curling around presumptive degenerating myelin ovoids within the CNS compartment of the dorsal root. In contrast, delaying NT-3 treatment in LES rats resulted in straighter, centrally-directed regenerating axons. These results indicate that regeneration may be best optimized through a combination of neurotrophin treatment plus complete clearance of myelin debris., Peer-reviewed article, Published. Received 3 June 2006, Revised 8 September 2006, Accepted 8 September 2006, Available online 22 November 2006.
Purification and characterization of a selective growth regulator for human myelopoietic progenitor cells
A monoclonal antibody, named CAMAL-1, was raised previously in our laboratory to a common antigen of acute myeloid leukemia (CAMAL), and was shown to be highly specific in its recognition of cells from patients with acute (AML) or chronic (CML) myelogenous leukemia. CAMAL was also reported to be prognostic of disease, in that patients whose numbers of CAMAL-1 reactive cells were high, or rose over time, had poorer prognoses and shorter survival times than patients whose CAMAL values were low or decreased. This correlation between CAMAL and disease prognosis led to the discovery that CAMAL-1immunoaffinity-purified leukemic cellular lysates contained a selective growth inhibitory activity for normal myeloid progenitor cells, since the growth of CML progenitors was not inhibited. The work described in this thesis focused primarily on the purification and characterization of the myelopoietic activity present in the CAMAL preparations, and its relationship to the leukemic marker (CAMAL). Initial purifications involved CAMAL-1immunoaffinity chromatography of leukemic cellular lysates, followed by FPLC molecular size fractionation and/or preparative SDS-PAGE. The myelopoietic activity was located within a30-35 kDa molecular weight fraction (P30), and the P30 fraction was consistently found to be selective in its inhibition of normal myeloid progenitors, since the growth of CML progenitors was not inhibited but was, in fact, stimulated. Antibodies were raised to P30 and used in the subsequent purification and characterization of the myelopoietic activity. Amino acid sequence analysis of the N-terminus and P30 tryptic peptides strongly suggested that P30 belonged to the serine protease family of enzymes, and the results obtained from protease assays indicated thatP30 preparations did possess enzyme activity. Prior to the completion of P30 molecular cloning experiments, however, the cDNA sequence for azurocidin/CAP37 was reported, and its predicted amino acid sequence was found to be identical to those obtained from the P30 protein samples. Azurocidin is a proteolytically inactive serine protease homologue, normally present in neutrophilic granules. Purifiedazurocidin did not possess inhibitory activity in normal progenitor cell assays; therefore, in order to isolate the biologic activity from azurocidin and other potentially contaminating proteins, P30 preparations were fractionated by reverse phase HPLC. The rpHPLC profiles were found to be similar to those reported for neutrophilic granules; however, the myelopoietica ctivity was obtained in a single rpHPLC fraction that aligned with the front portion of the azurocidin protein peak. Two dimensional isoelectric focusing/SDS-PAGE analysis of the biologically active rpHPLC fraction confirmed that it contained azurocidin, and no additional protein species were detected. Only the earlier eluting azurocidin rpHPLC fraction mediated the myelopoietic activity, and this fraction was also enriched in the higher molecular weight isoforms of azurocidin. Therefore, it appeared that a variably glycosylated isoform of azurocidin was mediating the biologic effects on myeloid progenitor cells, and because azurocidin obtained from normal neutrophils did not possess the myelopoietic activity, we speculate that the bioactive isoform of azurocidin is present in relatively higher amounts and/or is uniquely synthesized by leukemic cells., Thesis, Published.
Quantification of pyrrolizidine alkaloids in North American plants and honey by LC-MS
Pyrrolizidine alkaloids (PAs) are a class of naturally occurring compounds produced by many flowering plants around the World. Their presence as contaminants in food systems has become a significant concern in recent years. For example, PAs are often found as contaminants in honey through pollen transfer. A validated method was developed for the quantification of four pyrrolizidine alkaloids and one pyrrolizidine alkaloidN-oxide in plants and honey grown and produced in British Columbia. The method was optimised for extraction efficiency from the plant materials and then subjected to a single-laboratory validation to assess repeatability, accuracy, selectivity, LOD, LOQ and method linearity. The PA content in plants ranged from1.0 to 307.8 µg/g with repeatability precision between 3.8 and 20.8% RSD. HorRat values were within acceptable limits and ranged from 0.62 to 1.63 for plant material and 0.56–1.82 for honey samples. Method accuracy was determined through spike studies with recoveries ranging from 84.6 to 108.2% from the raw material negative control and from 82.1–106.0 % for the pyrrolizidine alkaloids in corn syrup. Based on the findings in this single-laboratory validation, this method is suitable for the quantitation of lycopsamine, senecionine, senecionineN-oxide, heliosupine and echimidine in common comfrey (Symphytum officinale), tansy ragwort (Senecio jacobaea), blueweed (Echium vulgare) and hound’s tongue (Cynoglossum officinale)and for PA quantitation in honey and found that PA contaminants were present at low levels in BC honey., Peer-reviewed article, Published. Received 4 June 2015; accepted 20 September 2015.

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