Is the carotenoid production from Phaffia rhodozyma yeast genuinely sustainable? a comprehensive analysis of biocompatibility, environmental assessment, and techno-economic constraints.

Microorganisms, such as yeasts, filamentous fungi, bacteria, and microalgae, have gained significant attention due to their potential in producing commercially valuable natural carotenoids. In recent years, Phaffia rhodozyma yeasts have emerged as intriguing non-conventional sources of carotenoids, particularly astaxanthin and ß-carotene. However, the shift from academic exploration to effective industrial implementation has been challenging to achieve. This study aims to bridge this gap by assessing various scenarios for carotenoid production and recovery. It explores the use of ionic liquids (ILs) and bio-based solvents (ethanol) to ensure safe extraction. The evaluation includes a comprehensive analysis involving Life Cycle Assessment (LCA), biocompatibility assessment, and Techno-Economic Analysis (TEA) of two integrated technologies that utilize choline-based ILs and ethanol (EtOH) for astaxanthin (+ß-carotene) recovery from P. rhodozyma cells. This work evaluates the potential sustainability of integrating these alternative solvents within a yeast-based bioeconomy.

Early Treatment with Pegylated Interferon Lambda for Covid-19.

BACKGROUND: The efficacy of a single dose of pegylated interferon lambda in preventing clinical events among outpatients with acute symptomatic coronavirus disease 2019 (Covid-19) is unclear. METHODS: We conducted a randomized, controlled, adaptive platform trial involving predominantly vaccinated adults with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Brazil and Canada. Outpatients who presented with an acute clinical condition consistent with Covid-19 within 7 days after the onset of symptoms received either pegylated interferon lambda (single subcutaneous injection, 180 µg) or placebo (single injection or oral). The primary composite outcome was hospitalization (or transfer to a tertiary hospital) or an emergency department visit (observation for >6 hours) due to Covid-19 within 28 days after randomization. RESULTS: A total of 933 patients were assigned to receive pegylated interferon lambda (2 were subsequently excluded owing to protocol deviations) and 1018 were assigned to receive placebo. Overall, 83% of the patients had been vaccinated, and during the trial, multiple SARS-CoV-2 variants had emerged. A total of 25 of 931 patients (2.7%) in the interferon group had a primary-outcome event, as compared with 57 of 1018 (5.6%) in the placebo group, a difference of 51% (relative risk, 0.49; 95% Bayesian credible interval, 0.30 to 0.76; posterior probability of superiority to placebo, >99.9%). Results were generally consistent in analyses of secondary outcomes, including time to hospitalization for Covid-19 (hazard ratio, 0.57; 95% Bayesian credible interval, 0.33 to 0.95) and Covid-19-related hospitalization or death (hazard ratio, 0.59; 95% Bayesian credible interval, 0.35 to 0.97). The effects were consistent across dominant variants and independent of vaccination status. Among patients with a high viral load at baseline, those who received pegylated interferon lambda had lower viral loads by day 7 than those who received placebo. The incidence of adverse events was similar in the two groups. CONCLUSIONS: Among predominantly vaccinated outpatients with Covid-19, the incidence of hospitalization or an emergency department visit (observation for >6 hours) was significantly lower among those who received a single dose of pegylated interferon lambda than among those who received placebo. (Funded by FastGrants and others; TOGETHER ClinicalTrials.gov number, NCT04727424.).

A comparative life cycle assessment of graphene nanoplatelets- and glass fibre-reinforced poly(propylene) composites for automotive applications.

The circulation of motor vehicles with an exhaust system is one of the leading causes of pollution. Due to the risk factor for human health and contribution to climate change, countries have been growing efforts to achieve a sustainable level of air quality and limit carbon dioxide (CO2) emissions in recent years. Hence, there has been significant interest in developing technological innovations such as alternative fuels and lighter thermoplastic composites for auto applications. Thermoplastic nanocomposites show substantial properties improvements, incorporating much lower nanofiller percentages than fibre-reinforced thermoplastic composites, allowing for a reduction in the weight of automotive parts (AP). For these motivations, this study presents a comparative life cycle assessment (LCA) of poly(propylene) (PP)-based graphene nanoplatelets (GnPs) nanocomposite and PP/glass fibre (GF) composite for automotive applications. The AP selected as the functional unit was the front end part of 1.5 tons weight car. The LCA included preparing raw materials, AP manufacturing, AP use, and AP end-of-life (EoL). Three different EoL scenarios were considered in this analysis. Several midpoint environmental impact indicators were evaluated. Overall, the results suggest that the different EoL scenarios do not affect the global environmental impact of both AP systems. The environmental impacts of the AP depend on the type of material. The AP of the PP-based GnPs nanocomposite exhibited a weight 28 % lower than the AP of the PP/GF composite. This lightweight resulted in energy and emissions savings, especially during the AP use stage, which was the stage with the most significant contribution in most environmental impact categories. Using nanocomposite reduced the AP environmental impact from 17 % to 75 %, depending on the impact category. The study suggested that substituting traditional composite with a new lighter nanocomposite can decrease the global environmental impacts caused by AP.

Combining eutectic solvents and food-grade silica to recover and stabilize anthocyanins from grape pomace.

Concentrated in the skins of red grapes are the anthocyanins, the primary colorants responsible for the fruits' reddish-purple color. These colorants are recognized for their significant antioxidant properties and potent nutraceutical and pharmaceutical ingredients. Nevertheless, their widespread use is compromised by the (i) need for more efficient yet sustainable downstream processes for their recovery and (ii) by the challenges imposed by their poor stability. In this work, these drawbacks were overcome by applying eutectic solvents and stabilizing agents. Besides, the anthocyanins were successfully loaded into a solid host material (approved in both food and pharmaceutical sectors) based on silicon dioxide (SiO2, loading capacity: 1extract:7silica m/m). Summing up, with the process developed, the extraction yield (21 mganthocyanins.gbiomass-1) and the stability (under 55, 75, and 95 °C) of the recovered anthocyanins were over three times better than with the conventional process. Finally, the raw materials and solvents were recycled, allowing an economical and environmentally friendly downstream process.

An eco-friendly approach for the recovery of astaxanthin and ß-carotene from Phaffia rhodozyma biomass using bio-based solvents.

There is a growing demand in the development of environmentally friendly technologies, based on the use of more biocompatible solvents for the recovery of natural bioactive compounds. In this work, the red yeast Phaffia rhodozyma biomass was used as a source of carotenoids to develop an integrative and efficient platform that promotes the recovery of astaxanthin and ß-carotene using bio-based solvents (BioSs). The extraction aptitude of pure BioSs was evaluated and compared with the conventional organic method. At this point, the influence of the BioSs molecular structures involved in the extraction procedures were also investigated. Overall, envisaging the industrial application of the process, an integrative platform was proposed for the recovery of astaxanthin/ß-carotene from P. rhodozyma biomass and the recycle of the BioSs. The life cycle assessment of the proposed technology using EtOH was evaluated, validating the sustainability of BioSs in the process with environmental impact reduction of 3-12%.

Diagnostic application of sensitive and specific phage-exposed epitopes for visceral leishmaniasis and human immunodeficiency virus coinfection.

The diagnosis of visceral leishmaniasis (VL) has improved with the search of novel antigens; however, their performance is limited when samples from VL/human immunodeficiency virus (HIV)-coinfected patients are tested. In this context, studies conducted to identify more suitable antigens to detect both VL and VL/HIC coinfection cases should be performed. In the current study, phage display was performed using serum samples from healthy subjects and VL, HIV-infected and VL/HIV-coinfected patients; aiming to identify novel phage-exposed epitopes to be evaluated with this diagnostic purpose. Nine non-repetitive and valid sequences were identified, synthetized and tested as peptides in enzyme-linked immunosorbent assay experiments. Results showed that three (Pep2, Pep3 and Pep4) peptides showed excellent performance to diagnose VL and VL/HIV coinfection, with 100% sensitivity and specificity values. The other peptides showed sensitivity varying from 50.9 to 80.0%, as well as specificity ranging from 60.0 to 95.6%. Pep2, Pep3 and Pep4 also showed a potential prognostic effect, since specific serological reactivity was significantly decreased after patient treatment. Bioinformatics assays indicated that Leishmania trypanothione reductase protein was predicted to contain these three conformational epitopes. In conclusion, data suggest that Pep2, Pep3 and Pep4 could be tested for the diagnosis of VL and VL/HIV coinfection.

Phosphoinositide-3 kinase gamma regulates caspase-1 activation and leukocyte recruitment in acute murine gout.

This study investigates the participation of PI3Kγ in the development of joint inflammation and dysfunction in an experimental model of acute gout in mice. Acute gout was induced by injection of monosodium urate (MSU) crystals into the tibiofemoral joint of mice. The involvement of PI3Kγ was evaluated using a selective inhibitor and mice deficient for PI3Kγ (PI3Kγ-/- ) or with loss of kinase activity. Neutrophils recovered from the inflamed joint were quantified and stained for phosphorylated Akt (pAkt) and production of reactive oxygen species (ROS). The adherence of leukocytes to the joint microvasculature was assessed by intravital microscopy and cleaved caspase-1 by Western blot. Injection of MSU crystals induced massive accumulation of neutrophils expressing phosphorylated Akt. In the absence of PI3Kγ, there was reduction of pAkt expression, chemokine production, and neutrophil recruitment. Genetic or pharmacological inhibition of PI3Kγ reduced the adherence of leukocytes to the joint microvasculature, even in joints with established inflammation. Neutrophils from PI3Kγ-/- mice produced less ROS than wild-type neutrophils. There was decreased joint damage and dysfunction in the absence of PI3Kγ. In addition, in the absence of PI3Kγ activity, there was reduction of cleaved caspase-1 and IL-1ß production in synovial tissue after injection of MSU crystals and leukotriene B4 . Our studies suggest that PI3Kγ is crucial for MSU crystal-induced acute joint inflammation. It is necessary for regulating caspase-1 activation and for mediating neutrophil migration and activation. Drugs that impair PI3Kγ function may be useful to control acute gout inflammation.

Leishmania infantum ß-Tubulin Identified by Reverse Engineering Technology through Phage Display Applied as Theranostic Marker for Human Visceral Leishmaniasis.

Two Leishmania infantum mimotopes (B10 and C01) identified by phage display showed to be antigenic and immunogenic for visceral (VL) and tegumentary (TL) leishmaniasis; however, their biological targets in the parasites have not been identified. The aim of the present study was to investigate the native antigens expressing both mimotopes, and to use them in distinct immunological assays. For this, a subtractive phage display technology was used, where a combinatorial library of single-chain variable fragments (scFv) was employed and the most reactive monoclonal antibodies for each target were captured, being the target antigens identified by mass spectrometry. Results in immunoblotting and immunoprecipitation assays showed that both monoclonal scFvs antibodies identified the ß-tubulin protein as the target antigen in L. infantum. To validate these findings, the recombinant protein was cloned, purified and tested for the serodiagnosis of human leishmaniasis, and its immunogenicity was evaluated in PBMC derived from healthy subjects and treated or untreated VL patients. Results showed high diagnostic efficacy, as well as the development of a specific Th1 immune response in the cell cultures, since higher IFN-γ and lower IL-10 production was found.

High-through identification of T cell-specific phage-exposed mimotopes using PBMCs from tegumentary leishmaniasis patients and their use as vaccine candidates against Leishmania amazonensis infection.

In the current study, phage-exposed mimotopes as targets against tegumentary leishmaniasis (TL) were selected by means of bio-panning cycles employing sera of TL patients and healthy subjects, besides the immune stimulation of peripheral blood mononuclear cells (PBMCs) collected from untreated and treated TL patients and healthy subjects. The clones were evaluated regarding their specific interferon-γ (IFN-γ) and interleukin-4 (IL-4) production in the in vitro cultures, and selectivity and specificity values were calculated, and those presenting the best results were selected for the in vivo experiments. Two clones, namely A4 and A8, were identified and used in immunization protocols from BALB/c mice to protect against Leishmania amazonensis infection. Results showed a polarized Th1 response generated after vaccination, being based on significantly higher levels of IFN-γ, IL-2, IL-12, tumour necrosis factor-α (TNF-α) and granulocyte-macrophage colony-stimulating factor (GM-CSF); which were associated with lower production of specific IL-4, IL-10 and immunoglobulin G1 (IgG1) antibodies. Vaccinated mice presented significant reductions in the parasite load in the infected tissue and distinct organs, when compared with controls. In conclusion, we presented a strategy to identify new mimotopes able to induce Th1 response in PBMCs from TL patients and healthy subjects, and that were successfully used to protect against L. amazonensis infection.

Chronic Intermittent Hypoxia-Induced Vascular Dysfunction in Rats is Reverted by N-Acetylcysteine Supplementation and Arginase Inhibition.

Chronic intermittent hypoxia (CIH), the main attribute of obstructive sleep apnea (OSA), produces oxidative stress, endothelial dysfunction, and hypertension. Nitric oxide (NO) plays a critical role in controlling the vasomotor tone. The NO level depends on the L-arginine level, which can be reduced by arginase enzymatic activity, and its reaction with the superoxide radical to produce peroxynitrite. Accordingly, we hypothesized whether a combination of an arginase inhibitor and an antioxidant may restore the endothelial function and reduced arterial blood pressure (BP) in CIH-induced hypertensive rats. Male Sprague-Dawley rats 200 g were exposed either to CIH (5% O2, 12 times/h 8 h/day) or sham condition for 35 days. BP was continuously measured by radio-telemetry in conscious animals. After 14 days, rats were treated with 2(S)-amino-6-boronohexanoic acid (ABH 400 µg/kg day, osmotic pump), N-acetylcysteine (NAC 100 mg/kg day, drinking water), or the combination of both drugs until day 35. At the end of the experiments, external carotid and femoral arteries were isolated to determine vasoactive contractile responses induced by KCL and acetylcholine (ACh) with wire-myography. CIH-induced hypertension (~8 mmHg) was reverted by ABH, NAC, and ABH/NAC administration. Carotid arteries from CIH-treated rats showed higher contraction induced by KCl (3.4 ± 0.4 vs. 2.4 ± 0.2 N/m2) and diminished vasorelaxation elicits by ACh compared to sham rats (12.8 ± 1.5 vs. 30.5 ± 4.6%). ABH reverted the increased contraction (2.5 ± 0.2 N/m2) and the reduced vasorelaxation induced by ACh in carotid arteries from CIH-rats (38.1 ± 4.9%). However, NAC failed to revert the enhanced vasocontraction (3.9 ± 0.6 N/m2) induced by KCl and the diminished ACh-induced vasorelaxation in carotid arteries (10.7 ± 0.8%). Femoral arteries from CIH rats showed an increased contractile response, an effect partially reverted by ABH, but completely reverted by NAC and ABH/NAC. The impaired endothelial-dependent relaxation in femoral arteries from CIH rats was reverted by ABH and ABH/NAC. In addition, ABH/NAC at high doses had no effect on liver and kidney gross morphology and biochemical parameters. Thus, although ABH, and NAC alone and the combination of ABH/NAC were able to normalize the elevated BP, only the combined treatment of ABH/NAC normalized the vascular reactivity and the systemic oxidative stress in CIH-treated rats.

Association between polymorphism of tumor necrosis factor alpha (tnfα) in the region -308 g/a with tinnitus in the elderly with a history of occupational noise exposure.

CONTEXT: Tinnitus is a common disorder that occurs frequently across all strata of population and has an important health concern and is often associated with different forms of the hearing loss of varying severity. AIMS: To investigate the association between the polymorphism of tumor necrosis factor alpha (TNFα) in the region -308 G/A with the susceptibility to tinnitus in individuals with the history of exposure to occupational noise. SETTINGS AND DESIGN: This was a cross-sectional study with a sample of 179 independent elderly people above 60 years of age. MATERIALS AND METHODS: Information on exposure to occupational noise was obtained by interviews. Audiological evaluation was performed using pure tone audiometry and genotyped through polymerase chain reaction by restriction fragment length polymorphism. STATISTICAL ANALYSIS USED: Data were analyzed using the chi-square test and the odds ratio (OR), with the significance level set at 5%. RESULTS: Among elderly with tinnitus (43.01%), 33.76% had a history of exposure to occupational noise. A statistically significant association was found between genotype frequencies of the TNFα gene in the -308 G/A region and the complaint of tinnitus (P = 0.04 and χ2 = 4.19). The elderly with the G allele were less likely to have tinnitus due to occupational noise exposure when compared to those carrying the A allele (OR = 2.74; 95% CI: 1.56-4.81; P < 0.0005). CONCLUSION: This study suggests an association between the TNFα with susceptibility to tinnitus in individuals with a history of exposure to occupational noise.

Morphoanatomical and physicochemical profile of Piper callosum: valuable assessment for its quality control

ABSTRACT Piper callosum Ruiz & Pav., Piperaceae, popularly known as “elixir-paregórico” and “matricá” in Brazil, is used in folk medicine to treat gonorrhea, general pain, and digestive disorders, and has repellent, astringent, diuretic, depurative, and haemostatic properties. Despite the fact that this plant is sold as a traditional phytotherapeutic product, we did not find reports on its quality control. We, therefore, performed macroscopic, microscopic, histochemical, and physicochemical analyses using standard methods to establish botanical authentication and purity degree parameters for leaves and stem of this species in two forms: medicinal plant and herbal drug. We observed the size, shape, color, texture, fracture surface and transection characteristics, leaf venation patterns, and calluses are valuable diagnostic characters to identify the herbal drugs when they are not ground or powdered. Since medicinal plants and herbal drugs did not differ anatomically, the following key anatomical characters for P. callosum can be used for diagnostic purposes of both types raw plant materials: epicuticular wax and cuticular flanges patterns; collenchyma features; fibers in the midrib; arrangement pattern of the vascular bundles of the midrib and petiole; shape of the midrib, leaf margin, petiole, and stem; occurrence of raphides; and morphology of the starch grains. Acid lipids, essential oils, oleoresins, steroids, tannins and flavonoids were histochemically identified. Total ash (leaves: 11.25%; stem: 5.25%), sulphated ash (leaves: 68.02%; stem: 12.50%), acid-insoluble ash (leaves: 2.82%; stem: 0.27%), moisture (leaves: 8.60%; stem: 6.10%), loss on drying (leaves: 11.08%; stem: 8.58%), and pH (leaves: 5.57, stem: 5.28) values were determined. The order of analyzed metal levels in leaf and stem herbal drugs was Al > V > Cu > Mn > Cr > Ni. Similar levels of Cd and Co and low levels of Hg were found. The results obtained can be used as quality control parameters for medicinal plants and herbal drugs of P. callosum.

Leishmania infantum mimotopes and a phage-ELISA assay as tools for a sensitive and specific serodiagnosis of human visceral leishmaniasis.

Serological methods used to diagnose visceral leishmaniasis (VL) are considered minimally invasive, but they present problems related with their sensitivity and/or specificity. In this study, a subtractive selection using the phage display technology against antibodies from healthy subjects living in endemic and non-endemic areas of disease, as well as from Chagas disease patients and those developing active VL, was developed. The aim of this study was to select bacteriophage-fused epitopes to be used in the serodiagnosis of human VL. Eight phage clones were selected after the bio-panning rounds, and their reactivity was evaluated in a phage-ELISA assay against a human serological panel. A wild-type clone and the recombinant K39-based immunochromatographic test were used as controls. In the results, it was shown that all clones showed an excellent performance to serologically identify VL patients, demonstrating the feasibility of the isolated phages for developing a specific and sensitive serodiagnosis of human VL.

New serological tools for improved diagnosis of human tegumentary leishmaniasis.

Human tegumentary leishmaniasis (HTL), characterized by skin ulcers that may spread and cause dreadful and massive tissue destruction of the nose and mouth, is considered a neglected tropical disease, and it is a serious threat to global health due to its continuous expansion, favored by the lifecycle of its causative organism that is maintained in domestic animal reservoirs and anthropophilic sand fly species. Serodiagnosis of HTL is a great challenge due to many biological factors, including hampered specificity and/or sensitivity. This investigation addresses the unmet need for new diagnostic markers of HTL, and describes a simple platform to improve the serodiagnosis. A constrained conformational phage display random peptide library combined with a magnetic microsphere-based subtraction strategy was used to identify ligands with potential diagnostic applications. Six clones were selected against IgG antibodies from HTL patients, characterized by sequencing and confirmed by a phage-ELISA using sera from patients developing visceral leishmaniasis (n=20), Chagas disease (n=10), mucosal (n=30) and cutaneous (n=20) leishmaniasis; as well as from healthy subjects living in endemic (n=20) and non-endemic (n=30) areas of leishmaniasis. A wild-type M13-phage clone and a soluble Leishmania antigenic extract were used as negative and positive controls, respectively. Three clones reached 100% sensitivity and specificity, without any cross-reactivity with sera from patients with leishmaniasis-related diseases. Briefly, we describe for the first time a set of serological markers based on three immunodominant mimotopes that showed 100% accuracy, and that could be used in a phage-ELISA assay for the HTL serodiagnosis.

Transmembrane TNF-α is sufficient for articular inflammation and hypernociception in a mouse model of gout.

Gout manifests as recurrent episodes of acute joint inflammation and pain due to the deposition of monosodium urate (MSU) crystals within the affected tissue in a process dependent on NLRP3 inflammasome activation. The synthesis, activation, and release of IL-1ß are crucial for MSU-induced inflammation. The current study evaluated the mechanism by which TNF-α contributed to MSU-induced inflammation. Male C57BL/6J or transgenic mice were used in this study and inflammation was induced by the injection of MSU crystals into the joint. TNF-α was markedly increased in the joint after the injection of MSU. There was inhibition in the infiltration of neutrophils, production of CXCL1 and IL-1ß, and decreased hypernociception in mice deficient for TNF-α or its receptors. Pharmacological blockade of TNF-α with Etanercept or pentoxyfylline produced similar results. Mechanistically, TNF-α blockade resulted in lower amounts of IL-1ß protein and pro-IL-1ß mRNA transcripts in joints. Gene-modified mice that express only transmembrane TNF-α had an inflammatory response similar to that of WT mice and blockade of soluble TNF-α (XPro™1595) did not decrease MSU-induced inflammation. In conclusion, TNF-α drives expression of pro-IL-1ß mRNA and IL-1ß protein in experimental gout and that its transmembrane form is sufficient to trigger MSU-induced inflammation in mice.

Protective effects of resveratrol on hepatotoxicity induced by isoniazid and rifampicin via SIRT1 modulation.

Acute liver injury was induced in male BALB/c mice by coadministering isoniazid and rifampicin. In this work, the effects of resveratrol (1) were investigated in the hepatotoxicity caused by isoniazid-rifampicin in mice. Compound 1 was administered 30 min prior to isoniazid-rifampicin. Serum biochemical tests, liver histopathological examination, oxidative stress, myeloperoxidase activity, cytokine production (TNF-α, IL-12p70, and IL-10), and mRNA expression of SIRT1-7 and PPAR-γ/PGC1-α were evaluated. The administration of 1 significantly decreased aspartate transaminase and alanine aminotransferase levels, myeloperoxidase activity, and cytokine levels. Furthermore, 1 reverted the decrease of catalase and glutathione activities and ameliorated the histopathological alterations associated with antituberculosis drugs. Modulation of SIRT1 and PPAR-γ/PGC1-α expression is likely involved in the protective effects of 1. The results presented herein show that 1 was able to largely prevent the hepatotoxicity induced by isoniazid and rifampicin in mice, mainly by modulating SIRT1 mRNA expression.

Banana peel as an adsorbent for removing atrazine and ametryne from waters.

The feasibility of using banana peel for removal of the pesticides atrazine and ametryne from river and treated waters has been demonstrated, allowing the design of an efficient, fast, and low-cost strategy for remediation of polluted waters. The conditions for removal of these pesticides in a laboratory scale were optimized as sample volume = 50 mL, banana mass = 3.0 g, stirring time = 40 min, and no pH adjustment necessary. KF(sor) values for atrazine and ametryne were evaluated as 35.8 and 54.1 µg g(-1) (µL mL(-1)) by using liquid scintillation spectrometry. Adsorption was also evaluated by LC-ESI-MS/MS. As quantification limits were 0.10 and 0.14 µg L(-1) for both pesticides, sample preconcentration was not needed. Linear analytical curves (up to 10 µg L(-1)), precise results (RSD < 4.5%), good recoveries (82.9-106.6%), and a > 90% removal efficiency were attained for both pesticides. Water samples collected near an intensively cultivated area were adequately remedied.

Multi-pumping flow systems: the potential of simplicity.

Nowadays, the trend towards more compact, smarter and simpler devices is generally recognized as one of the most challenging aspects in the development of analytical instrumentation. Modern flow-based procedures do not escape this tendency. The level of integration and automation and the operational functionality of Multi-pumping flow systems (MPFS) would, in most of the situations, meet this requirement. The essential elements of MPFS are multiple solenoid actuated micro-pumps strategically positioned in the flow manifold, which are accountable for solutions insertion, propelling and commutation, conditioning the establishment and subsequent detection of the reaction zone. Being the only active components of the flow manifold they provide a great operational simplicity and assure a straightforward run-time control of important analytical variables. Moreover, the reduction of active components minimizes the probability of occurrence of equipment failures, malfunctions or errors. The low size and low cost of solenoid micro-pumps make them ideal tools to build up compact environmentally friendly analytical systems, which are characterized by low solutions consumptions and the minimisation of hazardous waste generation. Furthermore, the reproducible pulsed flowing stream produced by micro-pumps actuation has proven to be a valuable feature regarding sample/reagent mixing and reaction zone homogenisation.

A multipumping flow system for in vitro screening of peroxynitrite scavengers.

Peroxynitrite anion is a reactive nitrogen species formed in vivo by the rapid, controlled diffusion reaction between nitric oxide and superoxide radicals. By reacting with several biological molecules, peroxynitrite may cause important cellular and tissue deleterious effects, which have been associated with many diseases. In this work, an automated flow-based procedure for the in vitro generation of peroxynitrite and subsequent screening of the scavenging activity of selected compounds is developed. This procedure involves a multipumping flow system (MPFS) and exploits the ability of compounds such as lipoic acid, dihydrolipoic acid, cysteine, reduced glutathione, oxidized glutathione, sulindac, and sulindac sulfone to inhibit the chemiluminescent reaction of luminol with peroxynitrite under physiological simulated conditions. Peroxynitrite was generated in the MPFS by the online reaction of acidified hydrogen peroxide with nitrite, followed by a subsequent stabilization by merging with a sodium hydroxide solution to rapidly quench the developing reaction. The pulsed flow and the timed synchronized insertion of sample and reagent solutions provided by the MPFS ensure the establishment of the reaction zone only inside the flow cell, thus allowing maximum chemiluminescence emission detection. The results obtained for the assayed compounds show that, with the exception of oxidized glutathione, all are highly potent scavengers of peroxynitrite at the studied concentrations.

A critical comparison of analytical flow systems exploiting streamlined and pulsed flows.

Multipumping (MPFS) and multicommuted (MCFS) flow systems relying on pulsed and laminar flows were critically compared. The mixing conditions and dispersion associated with both systems were evaluated by simulating the sample with bromocresol green. The molybdenum blue method for phosphate determination in soil extracts was also implemented in both flow systems. Furthermore, laser-induced fluorescence (LIF) was applied to visualize the dispersing sample; rhodamine B was used as the fluorescent species. The pulsed flow enhanced the mixing of the solutions involved, thus reducing reagent consumption (48 and 96 microl for MPFS and MCFS), and improving sampling rate (67 and 144 h(-1) for MCFS and MPFS). For phosphate determination, results obtained with both systems were precise (r.s.d. < 0.5%; n = 10) and accurate. Analyses of the absorbance vs time/space LIF plots revealed that exploitation of pulsed flow led to a pronounced radial dispersion and to a limited axial dispersion, typical aspects of turbulent flows.