High-resolution pore-water and solid-phase analyses of sediment cores GeoB6520-3 (Hydrate Hole) and GeoB6521-2 (Worm Hole) of the northern Congo Fan

The geochemical cycling of barium was investigated in sediments of pockmarks of the northern Congo Fan, characterized by surface and subsurface gas hydrates, chemosynthetic fauna, and authigenic carbonates. Two gravity cores retrieved from the so-called Hydrate Hole and Worm Hole pockmarks were examined using high-resolution pore-water and solid-phase analyses. The results indicate that, although gas hydrates in the study area are stable with respect to pressure and temperature, they are and have been subject to dissolution due to methane-undersaturated pore waters. The process significantly driving dissolution is the anaerobic oxidation of methane (AOM) above the shallowest hydrate-bearing sediment layer. It is suggested that episodic seep events temporarily increase the upward flux of methane, and induce hydrate formation close to the sediment surface. AOM establishes at a sediment depth where the upward flux of methane from the uppermost hydrate layer counterbalances the downward flux of seawater sulfate. After seepage ceases, AOM continues to consume methane at the sulfate/methane transition (SMT) above the hydrates, thereby driving the progressive dissolution of the hydrates "from above". As a result the SMT migrates downward, leaving behind enrichments of authigenic barite and carbonates that typically precipitate at this biogeochemical reaction front. Calculation of the time needed to produce the observed solid-phase barium enrichments above the present-day depths of the SMT served to track the net downward migration of the SMT and to estimate the total time of hydrate dissolution in the recovered sediments. Methane fluxes were higher, and the SMT was located closer to the sediment surface in the past at both sites. Active seepage and hydrate formation are inferred to have occurred only a few thousands of years ago at the Hydrate Hole site. By contrast, AOM-driven hydrate dissolution as a consequence of an overall net decrease in upward methane flux seems to have persisted for a considerably longer time at the Worm Hole site, amounting to a few tens of thousands of years.

Next generation e-learning platform: virtual laboratory simulations

Bioscience subjects require a significant amount of training in laboratory techniques to produce highly skilled science graduates. Many techniques which are currently used in diagnostic, research and industrial laboratories require expensive equipment for single users; examples of which include next generation sequencing, quantitative PCR, mass spectrometry and other analytical techniques. The cost of the machines, reagents and limited access frequently preclude undergraduate students from using such cutting edge techniques. In addition to cost and availability, the time taken for analytical runs on equipment such as High Performance Liquid Chromatography (HPLC) does not necessarily fit with the limitations of timetabling. Understanding the theory underlying these techniques without the accompanying practical classes can be unexciting for students. One alternative from wet laboratory provision is to use virtual simulations of such practical which enable students to see the machines and interact with them to generate data. The Faculty of Science and Technology at the University of Westminster has provided all second and third year undergraduate students with iPads so that these students all have access to a mobile device to assist with learning. We have purchased licences from Labster to access a range of virtual laboratory simulations. These virtual laboratories are fully equipped and require student responses to multiple answer questions in order to progress through the experiment. In a pilot study to look at the feasibility of the Labster virtual laboratory simulations with the iPad devices; second year Biological Science students (n=36) worked through the Labster HPLC simulation on iPads. The virtual HPLC simulation enabled students to optimise the conditions for the separation of drugs. Answers to Multiple choice questions were necessary to progress through the simulation, these focussed on the underlying principles of the HPLC technique. Following the virtual laboratory simulation students went to a real HPLC in the analytical suite in order to separate of asprin, caffeine and paracetamol. In a survey 100% of students (n=36) in this cohort agreed that the Labster virtual simulation had helped them to understand HPLC. In free text responses one student commented that "The terminology is very clear and I enjoyed using Labster very much”. One member of staff commented that “there was a very good knowledge interaction with the virtual practical”.

Bile And Liver Metallothionein Behavior In Copper-exposed Fish.

The present study analyzed metallothionein (MT) excretion from liver to bile in Nile Tilapia (Oreochromis niloticus) exposed to sub-lethal copper concentrations (2mgL(-1)) in a laboratory setting. MTs in liver and bile were quantified by spectrophotometry after thermal incubation and MT metal-binding profiles were characterized by size exclusion high performance liquid chromatography coupled to ICP-MS (SEC-HPLC-ICP-MS). Results show that liver MT is present in approximately 250-fold higher concentrations than bile MT in non-exposed fish. Differences between the MT profiles from the control and exposed group were observed for both matrices, indicating differential metal-binding behavior when comparing liver and bile MT. This is novel data regarding intra-organ MT comparisons, since differences between organs are usually present only with regard to quantification, not metal-binding behavior. Bile MT showed statistically significant differences between the control and exposed group, while the same did not occur with liver MT. This indicates that MTs synthesized in the liver accumulate more slowly than MTs excreted from liver to bile, since the same fish presented significantly higher MT levels in liver when compared to bile. We postulate that bile, although excreted in the intestine and partially reabsorbed by the same returning to the liver, may also release MT-bound metals more rapidly and efficiently, which may indicate an efficient detoxification route. Thus, we propose that the analysis of bile MTs to observe recent metal exposure may be more adequate than the analysis of liver MTs, since organism responses to metals are more quickly observed in bile, although further studies are necessary.

Supercritical Carbon Dioxide Extraction Of Capsaicinoids From Malagueta Pepper (capsicum Frutescens L.) Assisted By Ultrasound.

Extracts from malagueta pepper (Capsicum frutescens L.) were obtained using supercritical fluid extraction (SFE) assisted by ultrasound, with carbon dioxide as solvent at 15MPa and 40°C. The SFE global yield increased up to 77% when ultrasound waves were applied, and the best condition of ultrasound-assisted extraction was ultrasound power of 360W applied during 60min. Four capsaicinoids were identified in the extracts and quantified by high performance liquid chromatography. The use of ultrasonic waves did not influence significantly the capsaicinoid profiles and the phenolic content of the extracts. However, ultrasound has enhanced the SFE rate. A model based on the broken and intact cell concept was adequate to represent the extraction kinetics and estimate the mass transfer coefficients, which were increased with ultrasound. Images obtained by field emission scanning electron microscopy showed that the action of ultrasonic waves did not cause cracks on the cell wall surface. On the other hand, ultrasound promoted disturbances in the vegetable matrix, leading to the release of extractable material on the solid surface. The effects of ultrasound were more significant on SFE from larger solid particles.

Pharmacokinetic And Pharmacodynamic Evaluation Of A Nanotechnological Topical Formulation Of Lidocaine/prilocaine (nanorap) In Healthy Volunteers.

Nanorap is a new nanotechnological formulation for topical anesthesia composed of lidocaine (2.5%) and prilocaine (2.5%). The present study evaluated the pharmacokinetics (PK) of Nanorap. For the determination of lidocaine and prilocaine in human plasma a new method using high-performance liquid-chromatography coupled to tandem mass spectrometry was developed. Nanorap pharmacodynamic (PD) and its physical proprieties were also evaluated. Nanorap was administered by topical application of 2g to healthy volunteers and blood samples were collected for the PK analysis. The drugs were extracted from plasma by liquid-liquid extraction with ether/hexane (80/20, v/v). The chromatography separation was performed on a Genesis C18 analytical column 4 µm (100 x 2.1 mm i.d.) with a mobile phase of methanol/acetonitrile/water (40/30/30, for lidocaine, and 50/30/20, for prilocaine, v/v/v) + 2 mM of ammonium acetate and ropivacaine as internal standard. The drugs were quantified using a mass spectrometer with an electrospray source in the ESI positive mode (ES+) configured for multiple reaction monitoring. The PD of Nanorap was evaluated with the use of a visual analogue scale. Nanorap was characterized by cryofracture. The chromatography run time was 5.5 min for lidocaine and 3.3 min for prilocaine and the lower limit of quantification was 0.05 ng/mL for both drugs. Mean Cmax was 6.62 and 1.72 ng/mL for lidocaine and prilocaine, respectively. Median Tmax was 6.5 hours for both drugs. Nanocapsules had a mean size of 88nm and mean drug association of 92.5% and 89% for lidocaine and prilocaine, respectively. The PD study showed that Nanorap has a sufficient analgesic effect (>30% reduction in pain) after 10 minutes of application. A new simple, selective and sensitive method for determination of lidocaine and prilocaine in human plasma was developed. Nanorap generated safe plasma levels of the drugs and satisfactory analgesic effect.

In Vitro Biological Screening Of The Anticholinesterase And Antiproliferative Activities Of Medicinal Plants Belonging To Annonaceae.

The aim of this research was to investigate the antiproliferative and anticholinesterase activities of 11 extracts from 5 Annonaceae species in vitro. Antiproliferative activity was assessed using 10 human cancer cell lines. Thin-layer chromatography and a microplate assay were used to screen the extracts for acetylcholinesterase (AchE) inhibitors using Ellman's reagent. The chemical compositions of the active extracts were investigated using high performance liquid chromatography. Eleven extracts obtained from five Annonaceae plant species were active and were particularly effective against the UA251, NCI-470 lung, HT-29, NCI/ADR, and K-562 cell lines with growth inhibition (GI50) values of 0.04-0.06, 0.02-0.50, 0.01-0.12, 0.10-0.27, and 0.02-0.04 µg/mL, respectively. In addition, the Annona crassiflora and A. coriacea seed extracts were the most active among the tested extracts and the most effective against the tumor cell lines, with GI50 values below 8.90 µg/mL. The A. cacans extract displayed the lowest activity. Based on the microplate assay, the percent AchE inhibition of the extracts ranged from 12 to 52%, and the A. coriacea seed extract resulted in the greatest inhibition (52%). Caffeic acid, sinapic acid, and rutin were present at higher concentrations in the A. crassiflora seed samples. The A. coriacea seeds contained ferulic and sinapic acid. Overall, the results indicated that A. crassiflora and A. coriacea extracts have antiproliferative and anticholinesterase properties, which opens up new possibilities for alternative pharmacotherapy drugs.

Determination Of The Phenolic Composition From Brazilian Tropical Fruits By Uhplc-ms/ms.

Although Brazil is the third largest fruit producer in the world, several specimens consumed are not well studied from the chemical viewpoint, especially for quantitative analysis. For this reason and the crescent employment of mass spectrometry (MS) techniques in food science we selected twenty-two phenolic compounds with important biological activities and developed an ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method using electrospray (ESI) in negative ion mode aiming their quantification in largely consumed Brazilian fruits (açaí-do-Amazonas, acerola, cashew apple, camu-camu, pineapple and taperebá). Multiple reaction monitoring (MRM) was applied and the selection of proper product ions for each transition assured high selectivity. Linearity (0.99580%), precision (CV<20%) and extraction recovery rate (>80%) were satisfactory and showed that the method provides an efficient protocol to analyze phenolic compounds in fruit pulp extracts.

Development Of Egg Pc/cholesterol/α-tocopherol Liposomes With Ionic Gradients To Deliver Ropivacaine.

Ropivacaine (RVC) is an aminoamide local anesthetic widely used in surgical procedures. Studies with RVC encapsulated in liposomes and complexed in cyclodextrins have shown good results, but in order to use RVC for lengthy procedures and during the postoperative period, a still more prolonged anesthetic effect is required. This study therefore aimed to provide extended RVC release and increased upload using modified liposomes. Three types of vesicles were studied: (i) large multilamellar vesicle (LMV), (ii) large multivesicular vesicle (LMVV) and (iii) large unilamellar vesicle (LUV), prepared with egg phosphatidylcholine/cholesterol/α-tocopherol (4:3:0.07 mol%) at pH 7.4. Ionic gradient liposomes (inside: pH 5.5, pH 5.5 + (NH4)2SO4 and pH 7.4 + (NH4)2SO4) were prepared and showed improved RVC loading, compared to conventional liposomes (inside: pH 7.4). An high-performance liquid chromatography analytical method was validated for RVC quantification. The liposomes were characterized in terms of their size, zeta potential, polydispersion, morphology, RVC encapsulation efficiency (EE(%)) and in vitro RVC release. LMVV liposomes provided better performance than LMV or LUV. The best formulations were prepared using pH 5.5 (LMVV 5.5in) or pH 7.4 with 250 mM (NH4)2SO4 in the inner aqueous core (LMVV 7.4in + ammonium sulfate), enabling encapsulation of as much as 2% RVC, with high uptake (EE(%) ∼70%) and sustained release (∼25 h). The encapsulation of RVC in ionic gradient liposomes significantly extended the duration of release of the anesthetic, showing that this strategy could be a viable means of promoting longer-term anesthesia during surgical procedures and during the postoperative period.

Impact Of Drug Formulation And Free Platinum/cisplatin Ratio On Hypersensitivity Reactions To Cisplatin: Formulation Matters.

Use of cisplatin can induce type I hypersensitivity reactions that may also be linked to the quality of the drug utilized. We observed cases of hypersensitivity that appeared to be associated with the brand of cisplatin used. The aim of this study was to compare two different brands of cisplatin in relation to type I hypersensitivity reactions. Brand A was used in a tertiary care teaching hospital until 2012, and use of brand B started from January 2013, when the first hypersensitivity cases were observed. Patients were categorized based on symptom. Cisplatin of both brands was analysed by high-performance liquid chromatography (HPLC) and high-resolution electrospray ionization mass spectrometry (ESI-(+)-MS) and characterized according to US Pharmacopeia. There were no cases of hypersensitivity associated with the use of cisplatin brand A, whereas four of 127 outpatients that used cisplatin brand B were affected. The two brands were in accordance with the US Pharmacopeia parameters, and there was no significant difference in the total platinum levels between the two brands when analysed by HPLC. However, high-resolution ESI-(+)-MS analyses show that brand B contains approximately 2.7 times more hydrolysed cisplatin than brand A. The increase in the hydrolysed form of cisplatin found in brand B may be the cause of the hypersensitivity reaction observed in a subset of patients. We present the first study of the quality of drugs by high-resolution ESI-(+)-MS. Drug regulatory agencies and manufacturers should consider including measurement of hydrolysed cisplatin as a quality criterion for cisplatin formulations.

Fases estacionárias monolíticas para separações cromatográficas

Monolithic stationary phases represent a new generation of chromatographic separation media. These phases consist of a continuous separation bed prepared by in situ polymerization or consolidation inside the column tubing. In recent years, their simple preparation procedure, unique properties and excellent performance have attracted quite remarkable attention in liquid chromatography and capillary electrochromatography. This review summarizes the preparation, characterization and applications of monolithic stationary phases. The analytical potential of these columns is demonstrated with separations involving various families of compounds in different separation modes.

Validação e estimativa da incerteza de método para análise de licopeno e β-caroteno em polpa de tomate por cromatografia líquida de alta eficiência

A method to quantify lycopene and β-carotene in freeze dried tomato pulp by high performance liquid chromatography (HLPC) was validated according to the criteria of selectivity, sensitivity, precision and accuracy, and uncertainty estimation of measurement was determined with data obtained in the validation. The validated method presented is selective in terms of analysis, and it had a good precision and accuracy. Detection limit for lycopene and β-carotene was 4.2 and 0.23 mg 100 g-1, respectively. The estimation of expanded uncertainty (K = 2) for lycopene was 104 ± 21 mg 100 g-1 and for β-carotene was 6.4 ± 1.5 mg 100 g-1.

Aplicações de cromatografia líquida de alta eficiência para o estudo de poluentes orgânicos emergentes

Emerging organic pollutants (EOP) include many environmental contaminants based on commercial products such as pharmaceuticals, personal care products, detergents, gasoline, polymers, etc. EOP may be candidates for future regulation as they offer potential risk to environmental and human health due to their continual entrance into the environment and to the fact that even the most modern wastewater treatment plants are not able to totally transform / remove these compounds. High performance liquid chromatography is recommended to separate emerging organic pollutants with characteristics of high polarity and low volatility, especially pharmaceuticals, from environmental matrices.

Cromatografia líquida capilar: estado da arte e aplicações

This review describes the advantages and disadvantages of using capillary liquid chromatography (CLC), which is considered the newest member in the analytical separation science arsenal. Although CLC has tremendous potential for being the next major innovation in separatory analysis, it has not yet obtained great popularity compared to conventional high performance (and ultra-high performance) liquid chromatography. Comparisons are made between these techniques and some of the reasons that CLC has not yet reached its potential will be advanced.

Hidrocarbonetos aromáticos policíclicos em margarina, creme vegetal e maionese

Polycyclic aromatic hydrocarbons (PAHs) are a group of compounds that have been the subject of much concern due to their toxic potential. In this study, margarine?s, vegetable cream and mayonnaise available on the Brazilian market were analyzed for pyrene, chrysene, benzo(a)pyrene, benzo(b)fluoranthene and dibenzo(a,h)anthracene. The analytical methodology involved liquid-liquid extraction, clean-up on silica gel column and determination by high performance liquid chromatography using fluorescence detector. Variable levels of contamination were found within differents brands of the same product and within differents batches of the same brand. The total PAH content was in the range of 4.1 to 7.1mug/kg in vegetable cream, 1.7 to 3.9mug/kg in margarine and 1.0 to 21.7mug/kg in mayonnaise. In general the products which according to the label contain corn oil showed the highest levels of contamination. Based on these results and on the importance of fat, oils and derived products for the intake of PAHs, it is recommended that producers of margarine, vegetable creams and mayonnaise start to control the contamination of the vegetable oils used in the elaboration of these products, in order to reduce the exposure of consumers to excessive amounts of potentially carcinogenic compounds.

Metronidazole-containing gel for the treatment of periodontitis: an in vivo evaluation

The aim of this investigation was to monitor metronidazole concentrations in the gingival crevicular fluid (GCF) collected from periodontal pockets of dogs after treatment with an experimental 15% metronidazole gel. Five dogs had periodontitis induced by cotton ligatures placed subgingivally and maintained for a 30-day period. After the induction period, only pockets with 4 mm or deeper received the gel. Each pocket was filled up to the gingival margin by means of a syringe with a blunt-end needle. GCF was collected in paper strips and quantified in an electronic device before and after 15 minutes, 1 h, 6 h, 24 h and 48 h of gel administration. The GCF samples were assayed for metronidazole content by means of a high performance liquid chromatography method. Concentrations of metronidazole in the GCF of the 5 dogs (mean ± SD, in µg/mL) were 0 ± 0 before gel application and 47,185.75 ± 24,874.35 after 15 minutes, 26,457.34 ± 25,516.91 after 1 h, 24.18 ± 23.11 after 6 h, 3.78 ± 3.45 after 24 h and 3.34 ± 5.54 after 48 h. A single administration of the 15% metronidazole gel released the drug in the GCF of dogs in levels several-fold higher than the minimum inhibitory concentration for some periodontopathogens grown in subgingival biofilms for up to one hour, but metronidazole could be detected in the GCF at least 48 hours after the gel application.