Analysis of cannabinoids in oral fluid by liquid chromatography-tandem mass spectrometry

A sensitive method was developed for quantifying a wide range of cannabinoids in oral fluid (OF) by liquid chromatography-tandem mass spectrometry (LC-MS/MS). These cannabinoids include a dagger(9)-tetrahydrocannabinol (THC), 11-hydroxy-a dagger(9)-tetrahydrocannabinol (11-OH-THC), 11-nor-9-carboxy-a dagger(9)-tetrahydrocannabinol (THCCOOH), cannabinol (CBN), cannabidiol (CBD), a dagger(9)-tetrahydrocannabinolic acid A (THC-A), 11-nor-9-carboxy-a dagger(9)-tetrahydrocannabinol glucuronide (THCCOOH-gluc), and a dagger(9)-tetrahydrocannabinol glucuronide (THC-gluc). Samples were collected using a Quantisal (TM) device. The advantages of performing a liquid-liquid extraction (LLE) of KCl-saturated OF using heptane/ethyl acetate versus a solid-phase extraction (SPE) using HLB copolymer columns were determined. Chromatographic separation was achieved in 11.5 min on a Kinetex (TM) column packed with 2.6-mu m core-shell particles. Both positive (THC, 11-OH-THC, CBN, and CBD) and negative (THCCOOH, THC-gluc, THCCOOH-gluc, and THC-A) electrospray ionization modes were employed with multiple reaction monitoring using a high-end AB Sciex API 5000 (TM) triple quadrupole LC-MS/MS system. Unlike SPE, LLE failed to extract THC-gluc and THCCOOH-gluc. However, the LLE method was more sensitive for the detection of THCCOOH than the SPE method, wherein the limit of detection (LOD) and limit of quantification (LOQ) decreased from 100 to 50 pg/ml and from 500 to 80 pg/ml, respectively. The two extraction methods were successfully applied to OF samples collected from volunteers before and after they smoked a homemade cannabis joint. High levels of THC were measured soon after smoking, in addition to significant amounts of THC-A. Other cannabinoids were found in low concentrations. Glucuronide conjugate levels were lower than the method's LOD for most samples. Incubation studies suggest that glucuronides could be enzymatically degraded by glucuronidase prior to OF collection

Combining phospholipases and a liquid lipase for one-step biodiesel production using crude oils

Background Enzymatic biodiesel is becoming an increasingly popular topic in bioenergy literature because of its potential to overcome the problems posed by chemical processes. However, the high cost of the enzymatic process still remains the main drawback for its industrial application, mostly because of the high price of refined oils. Unfortunately, low cost substrates, such as crude soybean oil, often release a product that hardly accomplishes the final required biodiesel specifications and need an additional pretreatment for gums removal. In order to reduce costs and to make the enzymatic process more efficient, we developed an innovative system for enzymatic biodiesel production involving a combination of a lipase and two phospholipases. This allows performing the enzymatic degumming and transesterification in a single step, using crude soybean oil as feedstock, and converting part of the phospholipids into biodiesel. Since the two processes have never been studied together, an accurate analysis of the different reaction components and conditions was carried out. Results Crude soybean oil, used as low cost feedstock, is characterized by a high content of phospholipids (900 ppm of phosphorus). However, after the combined activity of different phospholipases and liquid lipase Callera Trans L, a complete transformation into fatty acid methyl esters (FAMEs >95%) and a good reduction of phosphorus (P <5 ppm) was achieved. The combination of enzymes allowed avoidance of the acid treatment required for gums removal, the consequent caustic neutralization, and the high temperature commonly used in degumming systems, making the overall process more eco-friendly and with higher yield. Once the conditions were established, the process was also tested with different vegetable oils with variable phosphorus contents. Conclusions Use of liquid lipase Callera Trans L in biodiesel production can provide numerous and sustainable benefits. Besides reducing the costs derived from enzyme immobilization, the lipase can be used in combination with other enzymes such as phospholipases for gums removal, thus allowing the use of much cheaper, non-refined oils. The possibility to perform degumming and transesterification in a single tank involves a great efficiency increase in the new era of enzymatic biodiesel production at industrial scale.

Subunit composition of seed storage proteins in high-protein soybean genotypes

The objective of this work was to quantify the accumulation of the major seed storage protein subunits, β-conglycinin and glycinin, and how they influence yield and protein and oil contents in high-protein soybean genotypes. The relative accumulation of subunits was calculated by scanning SDS-PAGE gels using densitometry. The protein content of the tested genotypes was higher than control cultivar in the same maturity group. Several genotypes with improved protein content and with unchanged yield or oil content were developed as a result of new breeding initiatives. This research confirmed that high-protein cultivars accumulate higher amounts of glycinin and β-conglycinin. Genotypes KO5427, KO5428, and KO5429, which accumulated lower quantities of all subunits of glycinin and β-conglycinin, were the only exceptions. Attention should be given to genotypes KO5314 and KO5317, which accumulated significantly higher amounts of both subunits of glycinin, and to genotypes KO5425, KO5319, KO539 and KO536, which accumulated significantly higher amounts of β-conglycinin subunits. These findings suggest that some of the tested genotypes could be beneficial in different breeding programs aimed at the production of agronomically viable plants, yielding high-protein seed with specific composition of storage proteins for specific food applications.

Simultaneous determination of gross alpha, gross beta and Ra-226 in natural water by liquid scintillation counting.

The determination of gross alpha, gross beta and 226Ra activity in natural waters is useful in a wide range of environmental studies. Furthermore, gross alpha and gross beta parameters are included in international legislation on the quality of drinking water [Council Directive 98/83/EC].1 In this work, a low-background liquid scintillation counter (Wallac, Quantulus 1220) was used to simultaneously determine gross alpha, gross beta and 226Ra activity in natural water samples. Sample preparation involved evaporation to remove 222Rn and its short-lived decay daughters. The evaporation process concentrated the sample ten-fold. Afterwards, a sample aliquot of 8 mL was mixed with 12 mL of Ultima Gold AB scintillation cocktail in low-diffusion vials. In this study, a theoretical mathematical model based on secular equilibrium conditions between 226Ra and its short-lived decay daughters is presented. The proposed model makes it possible to determine 226Ra activity from two measurements. These measurements also allow determining gross alpha and gross beta simultaneously. To validate the proposed model, spiked samples with different activity levels for each parameter were analysed. Additionally, to evaluate the model's applicability in natural water, eight natural water samples from different parts of Spain were analysed. The eight natural water samples were also characterised by alpha spectrometry for the naturally occurring isotopes of uranium (234U, 235U and 238U), radium (224Ra and 226Ra), 210Po and 232Th. The results for gross alpha and 226Ra activity were compared with alpha spectrometry characterization, and an acceptable concordance was obtained.

A Progress Report on Treating Loess, Fine Sands and Soft Limestones with Liquid Binders, HR-20, 1954

Certain areas of Iowa abound in loess, others contain soft limestones that are readily and cheaply available, and a large portion of the state is underlaid with sand. None of these materials is considered suitable in present practices for use in all-weather road construction. The loess is too fine and too difficult to handle; the limestones are considered too soft, and some of the harder ones unsound for this use; the sands are not naturally of the desired gradation and do not lend themselves to blending into satisfactory gradations. The purpose of this project is, therefore, to study and develop means and to determine the feasibility of using these materials, loess, fine sand, and soft limestones, either separately or in combinations in conjunction with liquid binders to produce paving mixtures applicable for all-weather road construction. Also included in the project was the development of methods of processing any of these materials, if necessary, to make them suitable for the desired purpose

Profiling of steroid metabolites after transdermal and oral administration of testosterone by ultra-high pressure liquid chromatography coupled to quadrupole time-of-flight mass spectrometry.

The screening of testosterone (T) misuse for doping control is based on the urinary steroid profile, including T, its precursors and metabolites. Modifications of individual levels and ratio between those metabolites are indicators of T misuse. In the context of screening analysis, the most discriminant criterion known to date is based on the T glucuronide (TG) to epitestosterone glucuronide (EG) ratio (TG/EG). Following the World Anti-Doping Agency (WADA) recommendations, there is suspicion of T misuse when the ratio reaches 4 or beyond. While this marker remains very sensitive and specific, it suffers from large inter-individual variability, with important influence of enzyme polymorphisms. Moreover, use of low dose or topical administration forms makes the screening of endogenous steroids difficult while the detection window no longer suits the doping habit. As reference limits are estimated on the basis of population studies, which encompass inter-individual and inter-ethnic variability, new strategies including individual threshold monitoring and alternative biomarkers were proposed to detect T misuse. The purpose of this study was to evaluate the potential of ultra-high pressure liquid chromatography (UHPLC) coupled with a new generation high resolution quadrupole time-of-flight mass spectrometer (QTOF-MS) to investigate the steroid metabolism after transdermal and oral T administration. An approach was developed to quantify 12 targeted urinary steroids as direct glucuro- and sulfo-conjugated metabolites, allowing the conservation of the phase II metabolism information, reflecting genetic and environmental influences. The UHPLC-QTOF-MS(E) platform was applied to clinical study samples from 19 healthy male volunteers, having different genotypes for the UGT2B17 enzyme responsible for the glucuroconjugation of T. Based on reference population ranges, none of the traditional markers of T misuse could detect doping after topical administration of T, while the detection window was short after oral TU ingestion. The detection ability of the 12 targeted steroids was thus evaluated by using individual thresholds following both transdermal and oral administration. Other relevant biomarkers and minor metabolites were studied for complementary information to the steroid profile, including sulfoconjugated analytes and hydroxy forms of glucuroconjugated metabolites. While sulfoconjugated steroids may provide helpful screening information for individuals with homozygotous UGT2B17 deletion, hydroxy-glucuroconjugated analytes could enhance the detection window of oral T undecanoate (TU) doping.

Health consultation : Rolfe aboveground storage tank petroleum bulk plant targeted brownfields assessment, Railroad Street and 300th Avenue, Rolfe, Pocahontas County, Iowa (2005)

The Iowa Department of Natural Resources (IDNR) has requested the Iowa Department of Public Health (IDPH) Hazardous Waste Site Health Assessment Program evaluate future health impacts of exposures at a former aboveground storage tank site located in Rolfe, Iowa. The former aboveground storage tank site is located to the southwest of the intersection of Railroad Street and 300th Avenue in Rolfe, Iowa. This site is undergoing a Targeted Brownfields Assessment conducted by the Contaminated Sites Section of the IDNR. This health consultation addresses potential health risks to people from future exposure to the soil within the property boundary, and any health impacts resulting from contaminated groundwater beneath the site property. The information in this health consultation was current at the time of writing. Data that emerges later could alter this document’s conclusions and recommendations.

Short-term storage in vitro and large-scale propagation of grapevine genotypes

The objective of this work was to evaluate the large-scale propagation of grapevine genotypes after short-term storage in vitro. Microshoots from ten grapevine genotypes were used. The following storage temperatures were evaluated: 10, 20, and 25°C. After short-term storage, the shoots were propagated in up to five successive subcultures, to assess the large-scale propagation of the germplasm maintained under conditions of minimal growth. The propagated shoots were rooted in different concentrations of indolbutiric acid (IBA) and acclimatized in greenhouse. The best temperature for short-term storage in vitro and survival of the genotypes was 20°C. In the propagation phase, the highest number of shoots per explant was found in the subcultures 4 and 5, with averages of 4.9 and 4.8 shoots per explant, respectively. In the rooting phase, the best results for number of roots were obtained using a culture medium supplemented with 0.4 µmol L-1 of IBA, with an average of three roots per shoot. During the acclimation phase, a survival rate higher than 95% was achieved after 30 days in the greenhouse. Grapevine genotypes maintained for six months in vitro, at 20ºC, can be micropropagated in large scale.

Quantifying sediment storage in a high Alpine valley (Turtmanntal, Switzerland)

The determination of sediment storage is a critical parameter in sediment budget analyses. But, in many sediment budget studies the quantification of magnitude and time-scale of sediment storage is still the weakest part and often relies on crude estimations only, especially in large drainage basins (>100km2). We present a new approach to storage quantification in a meso-scale alpine catchment of the Swiss Alps (Turtmann Valley, 110km2). The quantification of depositional volumes was performed by combining geophysical surveys and geographic information system (GIS) modelling techniques. Mean thickness values of each landform type calculated from these data was used to estimate the sediment volume in the hanging valleys and the trough slopes. Sediment volume of the remaining subsystems was determined by modelling an assumed parabolic bedrock surface using digital elevation model (DEM) data. A total sediment volume of 781·3×106?1005·7×106m3 is deposited in the Turtmann Valley. Over 60% of this volume is stored in the 13 hanging valleys. Moraine landforms contain over 60% of the deposits in the hanging valleys followed by sediment stored on slopes (20%) and rock glaciers (15%). For the first time, a detailed quantification of different storage types was achieved in a catchment of this size. Sediment volumes have been used to calculate mean denudation rates for the different processes ranging from 0·1 to 2·6mm/a based on a time span of 10ka. As the quantification approach includes a number of assumptions and various sources of error the values given represent the order of magnitude of sediment storage that has to be expected in a catchment of this size.

Liquid fructose down-regulates liver insulin receptor substrate 2 and gluconeogeneic enzymes by modifying nutrient sensing factors in rats

High consumption of fructose-sweetened beverages has been linked to a high prevalence of chronic metabolic diseases. We have previously shown that a short course of fructose supplementation as a liquid solution induces glucose intolerance in female rats. In the present work, we characterized the fructose-driven changes in the liver and the molecular pathways involved. To this end, female rats were supplemented or not with liquid fructose (10%, w/v) for 7 or 14 days. Glucose and pyruvate tolerance tests were performed, and the expression of genes related to insulin signaling, gluconeogenesis and nutrient sensing pathways was evaluated. Fructose-supplemented rats showed increased plasma glucose excursions in glucose and pyruvate tolerance tests and reduced hepatic expression of several genes related to insulin signaling, including insulin receptor substrate 2 (IRS-2). However, the expression of key gluconeogenic enzymes, glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, was reduced. These effects were caused by an inactivation of hepatic forkhead box O1 (FoxO1) due to an increase in its acetylation state driven by a reduced expression and activity of sirtuin 1 (SIRT1). Further contributing to FoxO1 inactivation, fructose consumption elevated liver expression of the spliced form of X-box-binding-protein-1 as a consequence of an increase in the activity of the mammalian target of rapamycin 1 and protein 38-mitogen activated protein kinase (p38-MAPK). Liquid fructose affects both insulin signaling (IRS-2 and FoxO1) and nutrient sensing pathways (p38-MAPK, mTOR and SIRT1), thus disrupting hepatic insulin signaling without increasing the expression of key gluconeogenic enzymes.

Influence of the pseudopotential on the properties of liquid rubidium at 315 K

The influence of the pseudopotential on both the structure and the self-diffusion of liquid rubidium at the melting point has been investigated by means of molecular-dynamics calculations. The model potential considered has been computed from the pseudopotential of Ashcroft, the dielectric function of Geldart and Vosko, and a Born-Mayer term. Four different values for the core radius which enters as input in the pseudopotential have been considered. In this way we have been able to observe and interpret the effect of this contribution on the properties of the liquid.

Storage protein profile and amino acid content in wild rice Oryza glumaepatula

- The objective of this work was to determine the total protein profile and the contents of the four major protein fractions (albumin, globulin, prolamin and glutelin) and of the amino acids in the endosperm of the rice wild species Oryza glumaepatula. The experiment was performed with 29 accessions of this species, collected from 13 Brazilian locations, and two commercial cultivars. Protein samples were prepared using dried, polished, and ground grains to obtain homogeneous, dry flour used in the preparation of extracts. Oryza glumaepatula accessions were identified with the highest levels of total protein, albumin and glutelin protein fractions, and amino acids (with the exception of tryptophan) in comparison to the two analized rice cultivars. The albumin and glutelin profiles in SDS-Page were distinct between rice cultivars and O. glumaepatula. This wild species has the potential to increase the nutritional quality of rice storage protein through interspecific crosses.

Shelf life and sensory assessment of tilapia quenelle during frozen storage

The objective of this work was to evaluate the shelf life and sensory attributes of tilapia quenelle. Treatments consisted of two types of packages - polyethylene zipper (retort pouch) (QA) and polyethylene waxed paper box (QB) - stored at -18ºC for 120 days. Tilapia quenelle was stable for all parameters established by Brazilian legislation. Average values of the evaluated attributes in different packages, during storage, showed no significant difference, except for the "refrigeration" flavor. However, during the storage period, there were significant differences for sensory attributes, as "moist appearance", fish and product aroma, and off flavors of "mud" and "refrigeration". Preserving product quality as for its sensory attributes, during storage, shows that tilapia quenelle is a convenience product and contributes to the increase of fish consumption.

Stocking densities and feeding strategies in shrimp and tilapia polyculture in tanks

The objective of this work was to evaluate the performance of Pacific marine shrimp (Litopenaeus vannamei) and tilapia (Oreochromis niloticus), in a polyculture in tanks subjected to different stocking densities and feeding strategies, in comparison with monoculture. Two experiments were performed, at the same time, in a completely randomized design with three treatments and four replicates each. Treatments for experiment I were: monoculture with 10 shrimp per m² (10S:0T); polyculture with 10 shrimp and 0.5 tilapia per m² (10S:0.5T); and polyculture with 10 shrimp and 1 tilapia per m² (10S:1T). Shrimp was the main crop, and feed was provided based on shrimp biomass. Treatments for experiment II were: monoculture with 2 tilapia per m² (2T:0S); polyculture with 2 tilapia and 2.5 shrimp per m² (2T:2.5S); and polyculture with 2 tilapia and 5 shrimp per m² (2T:5S). Tilapia was the main crop, and feed was provided based on fish requirements. In the experiment I, tilapia introduction to shrimp culture resulted in lower shrimp growth and poor feed conversion rate. In experiment II, shrimp introduction to tilapia culture did not interfere with fish performance. Polyculture is more efficient with the combination of 2 tilapia and 2.5 or 5 shrimp per m² and feed based on fish requirements.