Evaluation of Antipsychotic Drugs' Stability in Oral Fluid Samples.

Antipsychotics have narrow therapeutic windows, and their monitoring in biological fluids is therefore important; consequently, stability in those fluids must be investigated during method development and validation. This work evaluates the stability of chlorpromazine, levomepromazine, cyamemazine, clozapine, haloperidol, and quetiapine in oral fluid (OF) samples, using the dried saliva spots (DSS) sampling approach and gas chromatography coupled to tandem mass spectrometry. Since many parameters can influence the stability of the target analytes, design of experiments was adopted to check the crucial factors that affect that stability in a multivariate fashion. The studied parameters were the presence of preservatives at different concentrations, temperature, light, and time. It was possible to observe that antipsychotic stability improved when OF samples in DSS were stored at 4 °C, with a low ascorbic acid concentration, and in the absence of light. With these conditions, chlorpromazine and quetiapine were stable for 14 days, clozapine and haloperidol were stable for 28 days, levomepromazine remained stable for 44 days, and cyamemazine was stable for the entire monitored period (146 days). This is the first study that evaluates the stability of these antipsychotics in OF samples after application to DSS cards.

Automatic Identification of Myeloperoxidase Natural Inhibitors in Plant Extracts.

The aim of this study is the development of an automated method for myeloperoxidase activity evaluation and its application in testing the inhibitory action of different plant extracts on the activity of the enzyme. This enzyme has its concentration increased in inflammatory and infectious processes, so it is a possible target to limit these processes. Therefore, an automatic sequential in-jection analysis (SIA) system was optimized and demonstrated that it is possible to obtain results with satisfactory accuracy and precision. With the developed method, plant extracts were studied, as promising candidates for MPO inhibition. In the group of selected plant extracts, IC50 values from 0.029 ± 0.002 mg/mL to 35.4 ± 3.5 mg/mL were obtained. Arbutus unedo L. proved to be the most inhibitory extract for MPO based on its phenolic compound content. The coupling of an automatic SIA method to MPO inhibition assays is a good alternative to other conventional methods, due to its simplicity and speed. This work also supports the pharmacological use of these species that inhibit MPO, and exhibit activity that may be related to the treatment of infection and inflammation.

Essential oils used in dermocosmetics: Review about its biological activities.

INTRODUCTION: Currently, the demand for the use of constituents of natural origin in cosmetic formulations in detriment of synthetic compounds is noticeable. Several studies assess the potential of essential oils when incorporated into various cosmetic formulations and study their biological activities. This work intends to prepare a literature review on essential oils tested in dermocosmetic formulations and whose biological activities were evaluated through in vitro and/or in vivo tests. The main objectives for this study were as follows: to identify the essential oils that have been used in cosmetic formulations; and compile information on the main biological activities tested in cosmetic formulations. METHODS: A search was carried out until 2021 in the scientific databases PubMed and Web of Science, using different search terms, and several scientific articles from in vitro and in vivo studies in animals and clinical trials were selected and analyzed of involving development dermocosmetic formulations containing essential oils and the analysis of their biological activities. RESULTS: These studies demonstrate that the antimicrobial activity (antibacterial and antifungal) is the one most studied, mainly through in vitro tests. In vivo studies were also carried out either in animals or in clinical studies showing different effects, such as repellent action, inhibition of hair growth, and action against migraine. Regarding formulations, it was evident that creams are the most used. CONCLUSIONS: There is enormous potential for the use of essential oils in future formulations in the cosmetic and pharmaceutical industry, in particular as preservatives, exploring their other biological activities.

Solvent-Free Microwave Extraction of Thymus mastichina Essential Oil: Influence on Their Chemical Composition and on the Antioxidant and Antimicrobial Activities.

Solvent-free microwave extraction (SFME) is a combination of microwave heating and dry distillation performed at atmospheric pressure without the addition of water or organic solvents that has been proposed as a green method for the extraction of essential oils from aromatic and medicinal herbs. In this work, SFME and the conventional techniques of steam distillation (SD) and hydrodistillation (HD) were compared with respect to the extraction and antioxidant and antimicrobial activities of Thymus mastichina essential oil. The main constituent of essential oils obtained using different methods was 1,8-cineole (eucalyptol). The results showed that the essential oils extracted by means of SFME in 30 min were quantitatively (yield) and qualitatively (aromatic profile) similar to those obtained using conventional HD over 120 min. In addition, SFME generates less waste and less solvent, consumes less energy, and provides a higher yield for a shorter extraction time, which is advantageous for the extraction of the T. mastichina essential oil compared to SD. The antioxidant and antimicrobial activities of the T. mastichina essential oil obtained from either SFME or conventional extraction methods (SD or HD) showed a similar pattern. Large-scale experiments using this SFME procedure showed a potential industrial application.

Evaluation of Ionic Liquids and Ionic Liquids Active Pharmaceutical Ingredients Inhibition in Elastase Enzyme Activity.

Human neutrophil elastase (HNE) is used as diagnostic biomarker for inflammation/infection. In this work, 10 ionic liquids (ILs) and 11 ionic liquids active pharmaceutical ingredients (ILs-APIs) were tested to evaluate the inhibition effect on the activity of porcine pancreatic elastase enzyme, frequently employed as a model for HNE. The insertion of ionic liquids in some drugs is useful, as the insertion of ILs with inhibitory capacity will also slow down all processes in which this enzyme is involved. Therefore, a spectrophotometric method was performed to the determination of EC50 values of the compounds tested. EC50 values of 124 ± 4 mM to 289 ± 11 mM were obtained, with the most toxic IL for elastase being tetrabutylammonium acetate and the least toxic 1-butyl-3-methylimidazolium acetate. Moreover, sodium salicylate (raw material) presented the lower and benzethonium bistriflimide the higher EC50 when compared with all the IL-APIs tested. This work provides significant information about the effect of the studied IL and IL-APIs in elastase enzyme activity.

Allergic rhinitis characterization in community pharmacy customers: a cross-sectional study.

Background Allergic rhinitis represents a public health problem that is significantly prevalent in the global population and has been associated with asthma, a strong desire to sleep and a low quality of life. Objective This study aims to evaluate the prevalence, symptoms, control strategies and treatment, as well as the control of this condition and its impact on the quality of life of customers of community pharmacies with allergic rhinitis. Setting A questionnaire survey was carried out in nine community pharmacies in the city of Guarda, Portugal. Method In this cross-sectional study, data was collected by an interview between May 2014 and December 2014. The control of the illness and the impact of allergic rhinitis on the quality of life were assessed through a CARAT10 test and a WHOQOL-BREF instrument, respectively. Main outcome measure The impact of allergic rhinitis on the patient's quality of life. Results The estimated prevalence of allergic rhinitis was between 10.8% and 15.4%, from which 63 and 42 individuals were medically and symptomatically diagnosed, respectively, from a study population of 804 respondents. The majority of participants (57.1%) suffered from the symptoms more than twice a year. The symptoms, such as difficulty in falling asleep, repeated and continuous sneezing and bilateral nasal obstruction, were severe. There were patients with uncontrolled allergic rhinitis symptoms after the CARAT10 test, even when the individual's perception of the quality of life was good according to the WHOQOL-BREF score, with gender differences in the psychological domain. It should also be emphasized that there was a significant association between higher education levels with better control of the illness/quality of life. Additionally, most participants used pharmacological treatment (not alternative therapies) and the adoption of self-management measures to relieve their symptoms. Conclusions The findings of this study showed that the estimated prevalence of allergic rhinitis seems to be apparently lower in Guarda than that found in the general Portuguese population. From the data, some patients showed uncontrolled allergic rhinitis symptoms, strengthening the importance of the role of intervention by a health professional.

Thymus mastichina: Composition and Biological Properties with a Focus on Antimicrobial Activity.

Thymus mastichina has the appearance of a semishrub and can be found in jungles and rocky lands of the Iberian Peninsula. This work aimed to review and gather available scientific information on the composition and biological properties of T. mastichina. The main constituents of T. mastichina essential oil are 1,8-cineole (or eucalyptol) and linalool, while the extracts are characterized by the presence of flavonoids, phenolic acids, and terpenes. The essential oil and extracts of T. mastichina have demonstrated a wide diversity of biological activities. They showed antibacterial activity against several bacteria such as Escherichia coli, Proteus mirabilis, Salmonella subsp., methicillin-resistant and methicillin-sensitive Staphylococcus aureus, Listeria monocytogenes EGD, Bacillus cereus, and Pseudomonas, among others, and antifungal activity against Candida spp. and Fusarium spp. Additionally, it has antioxidant activity, which has been evaluated through different methods. Furthermore, other activities have also been studied, such as anticancer, antiviral, insecticidal, repellent, anti-Alzheimer, and anti-inflammatory activity. In conclusion, considering the biological activities reported for the essential oil and extracts of T. mastichina, its potential as a preservative agent could be explored to be used in the food, cosmetic, or pharmaceutical industries.

Xanthan Gum-Konjac Glucomannan Blend Hydrogel for Wound Healing.

Hydrogels are considered to be the most ideal materials for the production of wound dressings since they display a three-dimensional structure that mimics the native extracellular matrix of skin as well as a high-water content, which confers a moist environment at the wound site. Until now, different polymers have been used, alone or blended, for the production of hydrogels aimed for this biomedical application. From the best of our knowledge, the application of a xanthan gum-konjac glucomannan blend has not been used for the production of wound dressings. Herein, a thermo-reversible hydrogel composed of xanthan gum-konjac glucomannan (at different concentrations (1% and 2% w/v) and ratios (50/50 and 60/40)) was produced and characterized. The obtained data emphasize the excellent physicochemical and biological properties of the produced hydrogels, which are suitable for their future application as wound dressings.

Determination of opiates in whole blood using microextraction by packed sorbent and gas chromatography-tandem mass spectrometry.

In Portugal, and worldwide, the abuse of psychoactive substances is increasing, with a significant incidence of deaths related to their consumption. Opiates are one of the most prevalent group of substances in that context, and they are responsible for a significant impact of the mentioned harms. Therefore, it becomes necessary to equip labs with faster and effective methods to identify and quantify these substances. This work describes the development and validation of a novel analytical method for the simultaneous determination of morphine, codeine and 6-monoacetylmorphine in blood samples by gas chromatography-tandem mass spectrometry (GC-MS/MS), using microextraction by packed sorbent (MEPS) for sample preparation. Before the MEPS procedure, a precipitation step with acetonitrile was performed. The MEPS parameters were optimized using the fractional factorial planning (2k-1) and surface response methodology. The final optimized conditions were: number of strokes (20), amount of formic acid in the washing solution (3.36%), number of washes of the sorbent (1), amount of ammonium hydroxide in the elution solution (2.36%) and number of elution cycles (11). After the extraction procedure, the analytes were derivatized with MSTFA with 5% TMS. Using a sample volume of 250 µL, the method was validated according to internationally accepted standards. The method proved to be linear in the range of 5-1000 ng/mL with coefficients of determination greater than 0.99 for all analytes. Intra-and inter-day precision and accuracy were in accordance with the above-mentioned criteria, presenting coefficients of variation ≤15% and relative errors within a range of ± 15% of the theoretical concentration. The absolute recoveries ranged from 6 to 23%. The validated method was applied to the analysis of real samples, being an advantageous tool for the detection of those substances in blood. This is the first time that GCMS/MS with MEPS was used for the determination of these compounds in biological fluids.

Determination of methadone and EDDP in oral fluid using the dried saliva spots sampling approach and gas chromatography-tandem mass spectrometry.

The present work describes the development and validation of a novel approach to determine methadone (MTD) and its main metabolite (EDDP) in oral fluid samples, using the dried saliva spots (DSS) sampling approach and gas chromatography-tandem mass spectrometry (GC-MS/MS). Oral fluid samples (50 µL) were applied into Whatman™ 903 protein saver filter paper cards and were allowed to dry overnight. The extraction was carried out by immersion of the spot in 1 mL of isopropyl alcohol with agitation for 1 min. Afterwards, the extract was centrifuged for 15 min at 3500 rpm and the supernatant evaporated to dryness and reconstituted with 50 µL of methanol. The procedure was considered linear in the range of 10 to 250 ng/mL for both compounds, with determination coefficients greater than 0.99. Intra- and inter-day precision and accuracy revealed coefficients of variation (CVs) lower than 15% at the studied concentrations, with mean relative errors within ± 15% of the nominal concentrations. Recoveries ranged from 45 to 74%. The limits of detection and quantification were 5 and 10 ng/mL respectively for both analytes. All studied parameters complied with the defined criteria and the method enabled the successful determination of MTD and EDDP in oral fluid samples from patients undergoing opiate substitution/maintenance therapy.

Environmental Impact of Ionic Liquids: Automated Evaluation of the Chemical Oxygen Demand of Photochemically Degraded Compounds.

A novel automated fluorimetric technique was developed for the assessment of the chemical oxygen demand (COD) of ionic liquids (ILs) and combined with a photodegradation step to promote IL degradation. The method was implemented on a sequential injection analysis (SIA) system and is based on the reduction of cerium(IV) in the presence of irradiated ILs. Compounds incorporating the chloride anion were found to exhibit higher COD values and 1-butyl-3-methylimidazolium chloride ([bmim]+ [Cl]- ), 1-butyl-1-methylpyrrolidinium chloride ([bmpyr]+ [Cl]- ), and1-hexyl-3-methylimidazolium chloride ([hmim]+ [Cl]- ) also exhibited considerable photodegradability, whereas the cholinium cation and methanesulfonate and tetrafluoroborate anions showed resistance to photolysis. The developed methodology proved to be a simple, affordable, and robust method, showing good repeatability under the tested conditions (rsd <3.5 %, n=10). Therefore, it is expected that the developed approach can be used as a screening method for the preliminary evaluation of compounds' potential impact in the aquatic field. Additionally, the photolysis step presents an attractive option to promote degradation of ILs prior to their release into wastewater.

Assessment of ionic liquids' toxicity through the inhibition of acylase I activity on a microflow system.

Acylase I (ACY I) plays a role in the detoxication and bioactivation of xenobiotics as well in other physiological functions. In this context, an automated ACY I assay for the evaluation of ionic liquids' (ILs) toxicity was developed. The assay was implemented in a sequential injection analysis (SIA) system and was applied to eight commercially available ILs. The SIA methodology was based on the deacetylation of N-acetyl-l-methionine with production of l-methionine, which was determined using fluorescamine. ACY I inhibition in the presence of ILs was monitored by the decrease of fluorescence intensity. The obtained results confirmed the influence of ILs' structural elements on its toxicity and revealed that pyridinium and phosphonium cations, longer alkyl side chains and tetrafluoroborate anion displayed higher toxic effect on enzyme activity. The developed methodology proved to be robust and exhibited good repeatability (RSD < 1.3%, n = 10), leading also to a reduction of reagents consumption and effluents production. Thus, it is expected that the proposed assay can be used as a novel tool for ILs' toxicity screening.

Nanoparticle-based assays in automated flow systems: A review.

Nanoparticles (NPs) exhibit a number of distinctive and entrancing properties that explain their ever increasing application in analytical chemistry, mainly as chemosensors, signaling tags, catalysts, analytical signal enhancers, reactive species generators, analyte recognition and scavenging/separation entities. The prospect of associating NPs with automated flow-based analytical is undoubtedly a challenging perspective as it would permit confined, cost-effective and reliable analysis, within a shorter timeframe, while exploiting the features of NPs. This article aims at examining state-of-the-art on continuous flow analysis and microfluidic approaches involving NPs such as noble metals (gold and silver), magnetic materials, carbon, silica or quantum dots. Emphasis is devoted to NP format, main practical achievements and fields of application. In this context, the functionalization of NPs with distinct chemical species and ligands is debated in what concerns the motivations and strengths of developed approaches. The utilization of NPs to improve detector's performance in electrochemical application is out of the scope of this review. The works discussed in this review were published in the period of time comprised between the years 2000 and 2013.

Automatic flow methodology for kinetic and inhibition studies of reactions with poorly water-soluble substrates in ionic liquid systems.

In the present work an automatic generic tool, based on sequential injection analysis (SIA) for kinetic and inhibition studies of reactions with poorly water-soluble compounds in ionic liquid (IL)-containing systems, is described. The oxidation of the poorly water-soluble phenolic compound, caffeic acid, catalyzed by the mushroom tyrosinase, in different 1-butyl-3-methylimidazolium tetrafluoroborate ([bmim][BF(4)])/buffer mixtures as reaction media, was investigated. This determination was based on measuring depletion rate of the substrate caffeic acid at its maximum wavelength (λ(max) 311 nm). The influence of several parameters such as substrate and enzyme concentration, temperature, pH, delay times and measurement periods on the sensitivity and performance of the SIA system were studied and the optimum reaction conditions subsequently selected. The obtained results showed that tyrosinase was active in oxidising caffeic acid in this water-miscible IL and the presence of an impaired tyrosinase activity with increase in [bmim][BF(4)] concentration as an increase in the apparent Michaelis-Menten constant (K(M)(app)) was observed while the maximum reaction rate (V(max)(app)) remained fairly constant. The results were compared to those obtained when the assay was performed in water/methanol mixtures under the same conditions to substantiate [bmim][BF(4)] as an alternative to conventional organic solvents. Additionally, it was shown that tyrosinase is effectively inhibited by the substrate analogues tested (trans-cinnamic acid and 3,4-dihydroxybenzoic acid) in the IL-containing aqueous system used.

Flow system for the automatic screening of the effect of phenolic compounds on the luminol-hydrogen peroxide-peroxidase chemiluminescence system.

In this work, an automated flow-based procedure for the screening of the effect of the different phenolic compounds on the chemiluminescence (CL) luminol-hydrogen peroxide-horseradish peroxidase (HRP) system is presented. This procedure involves the combination of multisyringe flow injection analysis (MFSIA) and sequential injection analysis (SIA) techniques and exploits the ability of the different subgroups of phenols, such as cholorophenols, nitrophenols, methylphenols and polyphenols, to enhance or inhibit the described CL system. The implementation of this reaction in the SIA-MSFIA system enabled favourable and precise conditions to evaluate the effect of phenolic compounds, as it involves an in-line reaction between the phenolic derivative, hydrogen peroxide and peroxidase and subsequent oxidized HRP intermediates generation prior to the fast reaction with the chemiluminogenic reagent. Several studies were then performed with the aim of establishing the appropriate flow system configuration and reaction conditions. It was shown that phenol and chlorophenols produce an enhanced CL response and nitrophenols, methylphenols and polyphenols are inhibitors within the range of concentrations studied (1-100 mg/L). Based on these studies, the developed method was applied to the determination of total polyphenol and phenol content in wine/grape seeds and water samples, respectively, and the results obtained showed good agreement with those furnished by the corresponding Folin-Ciocalteu and 4-aminoantipyrine reference methods. The developed approach is further pursued by designing an automated generic tool for performing studies of peroxidase-catalysed CL reactions of luminol focused on the detection of compounds that will affect the rate of those reactions.

Sequential injection system for phospholipase A2 activity evaluation: studies on liposomes using an environment-sensitive fluorescent probe.

This work reports the development of an automatic methodology based on the use of 1-anilinonaphthalene-8-sulfonate (ANS) as an interfacial fluorescent probe for detecting the hydrophobic environment shift around the probe, caused by the hydrolytic action of PLA(2) on the liposomes. The implementation of this reaction in a sequential injection analysis (SIA) system along with the use of the mixing chambers permitted the evaluation of PLA(2) activity and assessment of the inhibitory effect of the non-steroidal anti-inflammatory drugs (NSAIDs) on PLA(2) activity. Several studies were performed with the aim of establishing the appropriate flow system configuration: the liposome substrate; PLA(2) and ANS optimum concentrations and incubation times before and after the enzyme addition. Based on these studies, the optimum reaction conditions were selected. It was shown that PLA(2) is effectively inhibited by the NSAIDs tested (meloxicam, tolmetin and ibuprofen) and by the alpha-lipoic acid, used as a positive control. Results obtained from the flow system are in agreement with those provided by the comparison batch procedures. The proposed methodology is in fact more efficient and rapid than the comparison batch experiments, enabling the exact timing of fluidic manipulations and precise control of the reaction conditions.

Flow methodology for methanol determination in biodiesel exploiting membrane-based extraction.

A methodology based in flow analysis and membrane-based extraction has been applied to the determination of methanol in biodiesel samples. A hydrophilic membrane was used to perform the liquid-liquid extraction in the system with the organic sample fed to the donor side of the membrane and the methanol transfer to an aqueous acceptor buffer solution. The quantification of the methanol was then achieved in aqueous solution by the combined use of immobilised alcohol oxidase (AOD), soluble peroxidase and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). The optimization of parameters such as the type of membrane, the groove volume and configuration of the membrane unit, the appropriate organic solvent, sample injection volume, as well as immobilised packed AOD reactor was performed. Two dynamic analytical working ranges were achieved, up to 0.015% and up to 0.200% (m/m) methanol concentrations, just by changing the volume of acceptor aqueous solution. Detection limits of 0.0002% (m/m) and 0.007% (m/m) methanol were estimated, respectively. The decision limit (CCalpha) and the detection capacity (CCbeta) were 0.206 and 0.211% (m/m), respectively. The developed methodology showed good precision, with a relative standard deviation (R.S.D.) <5.0% (n=10). Biodiesel samples from different sources were then directly analyzed without any sample pre-treatment. Statistical evaluation showed good compliance, for a 95% confidence level, between the results obtained with the flow system and those furnished by the gas chromatography reference method. The proposed methodology turns out to be more environmental friendly and cost-effective than the reference method.

Determination of total and oxidized glutathione in human whole blood with a sequential injection analysis system.

This work reports the development of a simple, robust, automated sequential injection analysis (SIA) system for the enzymatic determination of total (tGSH) and oxidized (GSSG) glutathione in human whole blood. The reduced (GSH) glutathione concentration is then obtained as the difference between the tGSH and GSSG concentrations. The determination was based on the DTNB-GSSG reductase recycling assay, which couples the specificity of the GSSG reductase (GR) with an amplification of the response to glutathione, followed by spectrophotometric detection of the 2-nitro-5-thiobenzoic acid (TNB) formed (lambda=412 nm). The implementation of this reaction in a SIA flow system with an in-line dilution strategy permitted the necessary distinct application ranges for tGSH and for GSSG. It also guaranteed the exact timing of fluidic manipulations and precise control of the reaction conditions. The influence of parameters such as reagents concentration, temperature, pH, flow rate of the carrier buffer solution, as well as reaction coil length, etc., on the sensitivity and performance of the SIA system were studied and the optimum reaction conditions subsequently selected. Linear calibration plots were obtained for GSH and GSSG concentrations up to 3.00 and 1.50 microM, with detection limits of 0.031 and 0.014 microM, respectively. The developed methodology showed good precision, with a relative standard deviation (R.S.D.)<5.0% (n=10) for determination of both glutathione forms. Statistical evaluation showed good compliance, for a 95% confidence level, between the results obtained with the SIA system and those furnished by the comparison batch procedure.