Development of an HPLC-MS/MS Method for Chiral Separation and Quantitation of (R)- and (S)-Salbutamol and Their Sulfoconjugated Metabolites in Urine to Investigate Stereoselective Sulfonation.

The aim of this study was to develop and optimize a chiral HPLC-MS/MS method for quantitative analysis of (R)-/(S)-salbutamol and (R)-/(S)-salbutamol-4'-O-sulfate in human urine to allow for bioanalytical quantitation of the targeted analytes and investigations of stereoselectivity in the sulfonation pathway of human phase Ⅱ metabolism. For analytical method development, a systematic screening of columns and mobile phases to develop a separation via enantiomerically selective high performance liquid chromatography was performed. Electrospray ionization settings were optimized via multiple-step screening and a full factorial design-of-experiment. Both approaches were performed matrix-assisted and the predicted values were compared. The full factorial design was superior in terms of prediction power and knowledge generation. Performing a longitudinal excretion study in one healthy volunteer allowed for the calculation of excretion rates for all four targeted analytes. For this proof-of-concept, either racemic salbutamol or enantiopure levosalbutamol was administered perorally or via inhalation, respectively. A strong preference for sulfonation of (R)-salbutamol for inhalation and peroral application was found in in vivo experiments. In previous studies phenol sulfotransferase 1A3 was described to be mainly responsible for salbutamol sulfonation in humans. Thus, in vitro and in silico investigations of the stereoselectivity of sulfotransferase 1A3 complemented the study and confirmed these findings.

Deep eutectic solvents combined with beta-cyclodextrin derivatives for chiral separation of typical adrenergic receptor agonists by capillary electrophoresis with amperometric detection.

Enantioseparation has always been one of the research hotspots and difficulties in the field of modern separation science. In this work, a binary chiral electrophoretic separation system was constructed using deep eutectic solvents (DESs) coupled with beta-cyclodextrin derivatives based on capillary electrophoresis with amperometric detection system, and five groups of typical adrenergic receptor agonists (adrenaline, salbutamol, isoproterenol, norepinephrine and terbutaline) were selected as the model enantiomers. The effects of additive types and contents of DESs and cyclodextrins, and the pH value and concentration of the running buffer on the resolution of the selected chiral compounds were investigated in detail. The mechanism of DESs improving separation was explored preliminarily by means of UV spectrophotometry, which was further verified based on the comparison of single and mixed components of choline chlorine-urea DES. Under the optimum conditions, the relative standard deviations for inter-day and intra-day repeatability of the migration time, peak area and resolution for adrenaline and salbutamol were within 8.7%, and the limits of detection reached 0.030 µg mL-1 (S/N = 3). The recovery data were in the range of 96.3-118.7%. The developed methods have been applied for the analyses of (+)-adrenaline hydrochloride injection and (±)-salbutamol aerosol. This binary chiral electrophoretic separation system by CE-AD has high detection sensitivity and low analytical cost, providing an alternative for the separation and analysis of chiral drugs.

Immunochromatographic Assay Based on Polydopamine-Decorated Iridium Oxide Nanoparticles for the Rapid Detection of Salbutamol in Food Samples.

Salbutamol (SAL), a ß-2 adrenoreceptor agonist, is an unpopular addition to livestock and poultry, causing several side effects to human health. Thus, it is very important to develop a simple and rapid analytical method to screen SAL in the field of food safety. Here, we present an immunochromatographic assay (ICA) method for sensitively detecting SAL with polydopamine-decorated iridium oxide nanoparticles (IrO2@PDA NPs) as a signal tag. The IrO2@PDA with excellent hydrophilicity, biocompatibility, and stability was synthesized by oxidating self-polymerization of dopamine hydrochloride (DAH) on the surface of IrO2 NPs and used to label monoclonal antibodies (mAbs) through simple physical adsorption. Compared with IrO2 NPs, the IrO2@PDA also possessed superior optical properties and higher affinity with mAbs. With the proposed method, the limit of detection for SAL was 0.002 ng/mL, which was improved at least 24-fold and 180-fold compared with the IrO2 NPs-based ICA and conventional gold nanoparticles-based ICA, respectively. Furthermore, the SAL residuals in pork, pork liver, and beef were successfully detected by the developed biosensor and the recoveries ranged from 85.56% to 115.56%. Briefly, this work indicated that the powerful IrO2@PDA-based ICA can significantly improve detection sensitivity and has huge potential for accurate and sensitive detection of harmful small molecules analytes in food safety fields.

High-Performance Liquid Chromatography Method for Simultaneous Determination of Guaifenesin, Salbutamol Sulfate and Guaifenesin Impurity (Guaiacol).

A simple and reliable HPLC method is developed for isocratic separation of a ternary mixture of Salbutamol Sulfate (SAL), Guaifenesin (GUI) and its impurity Guaiacol (GUA) either in pure powder or in pharmaceutical formulation. Chromatographic separation was applied on a Hypersil GOLD CN column with a mobile phase consisting of 0.05 M KH2PO4 (containing 0.1% triethylamine, pH adjusted to 3.7 by phosphoric acid): methanol (60: 40 by volume) using 0.6 mL min-1 flow rate and detection of peaks at 275 nm at 25°C with run time around 6 min. The calibration plots were linear over the concentration ranges of 0.5-20, 0.5-30 and 0.1-10 µg mL-1 for SAL, GUI and GUA, respectively. ICH guidelines were used for the validation of presented method. The obtained results were compared with the results obtained from the reported HPLC method and no significant difference was found regarding accuracy and precision.

An in vitro investigation into the release of fugitive medical aerosols into the environment during manual ventilation.

BACKGROUND: During manual resuscitation, nebulizer therapy may be used to deliver therapeutics to patients in respiratory distress. However, the devices used to generate and deliver these medical aerosols have the potential to release these therapeutics into the local environment and expose caregivers to unwanted medical aerosols. AIM: To quantify the levels of fugitive medical aerosol released into the environment during aerosol drug delivery using a manual resuscitation bag with and without filtration. METHODS: Time-varying fugitive aerosol concentrations were measured using an aerodynamic particle sizer placed at a position designed to mimic a caregiver. Two nebulizer types were assessed, a vibrating mesh nebulizer and a jet nebulizer. The aerosol dose delivered to the simulated patient lung was also quantified. FINDINGS: Filtration of the exhalation port of the manual resuscitation bag was seen to reduce fugitive medical aerosols to ambient levels for both nebulizer types. The vibrating mesh nebulizer delivered the greatest quantity of aerosol to the simulated adult patient (18.44 ± 1.03% versus 3.64 ± 0.26% with a jet nebulizer). CONCLUSIONS: The results highlight the potential for exposure to fugitive medical aerosols released during the delivery of aerosol therapy with a manual resuscitation bag and also the potential for significant variation in patient lung dose depending on nebulizer type.

Pharmacokinetics of salmeterol and its main metabolite α-hydroxysalmeterol after acute and chronic dry powder inhalation in exercising endurance-trained men: Implications for doping control.

As of 2020, use of beta2 -agonist salmeterol is restricted by the World Anti-Doping Agency (WADA) and is only permitted by inhalation at therapeutic doses not exceeding 200 µg in 24 h. In contrast to beta2 -agonists salbutamol and formoterol, WADA has not established a urine threshold for salmeterol despite its muscle hypertrophic actions observed in animals. Herein, we investigated plasma (0-4 h) and urine (0-24 h) concentrations (by ultra-high-performance liquid chromatography-tandem mass spectrometry [UHPLC-MS/MS]) of salmeterol and α-hydroxysalmeterol after dry powder inhalation at supratherapeutic (400 µg) and high therapeutic (200 µg) doses, and after seven consecutive days of therapeutic inhalation (200 µg × day-1 ) in 11 healthy endurance-trained men. During each trial, participants inhaled salmeterol before 1½ h moderate-intensity cycling. Mean ± SD maximum urine concentrations of salmeterol unadjusted for specific gravity reached 4.0 ± 1.6, 2.1 ± 1.5, and 2.2 ± 1.1 ng × ml-1 for 400 µg, 200 µg, and seven consecutive days of 200 µg, respectively, with corresponding maximum urine concentrations of α-hydroxysalmeterol being 11.6 ± 6.1, 5.7 ± 4.6, and 6.5 ± 2.6 ng × ml-1 . Within the relevant window for doping control (first 6 h post-inhalation), the present data (119 samples), along with 64 biobank urine samples, showed that a combined salmeterol and α-hydroxysalmeterol urine threshold with equal cut-offs of 3.3 ng × ml-1 was superior to a salmeterol-only threshold to discriminate therapeutic (200 µg) from supratherapeutic use (400 µg) with a sensitivity of 24% with 0% false positives when applying the WADA technical document (TD2019DL.v2) method of specific gravity adjustment. Thus, a combination of urine salmeterol and α-hydroxysalmeterol concentrations may be suitable for discriminating between therapeutic and supratherapeutic prohibited inhalation of salmeterol.

Bilinear and trilinear algorithms utilizing full and selected variables for resolution and quantitation of four components with overlapped spectral signals in bulk and syrup dosage form.

Spectrophotometric-assisted chemometric techniques are beneficial for resolving spectral overlapping and are considered comparable to traditional chromatographic methods. In this work, different chemometric approaches were applied for simultaneous determination of Bromhexine HCl (BRHX), Guaifenesin (GUA) and Salbutamol sulphate (SALB) in the presence of Guaiacol (GUAIA), without any prior separation. Two-way and three-way techniques were applied. The resolving power of genetic algorithm (GA-PLS), trilinear partial least square (N-PLS) and multivariate curve resolution (MCR-ALS) were investigated. A set of 17 synthetic samples in the concentration range 10.0-30.0 µg/mL of BRHX, GUA and SALB and 6.0-10.0 µg/mL of GUAIA were used in the construction of the calibration models. Commercially available syrup dosage form was successfully analyzed by the developed methods without interference from formulation additives. The developed models were evaluated through calculation of root mean squared error of prediction (RMSEP), the obtained values were 0.263, 0.419 and 0.342 for BRHX, 0.254, 0.318 and 0.503 for GUA and 0.298, 0.268 and 0.302 for SALB using N-PLS, MCR-ALS and GA-PLS, respectively. The resolving power of the developed models was emphasized through comparison with a reported HPLC method, where no significant difference was found regarding both accuracy and precision.

Development of a highly sensitive time-resolved fluoroimmunoassay for the determination of trace salbutamol in environmental samples.

Veterinary drug residues have become a major source of environmental pollutants. To monitor trace salbutamol (SAL) in the environment, a highly sensitive and reliable time-resolved fluoroimmunoassay (TRFIA) was developed. Under the optimum parameters, the half-maximal inhibition concentration (IC50) and the limit of detection (LOD, IC10) were determined to be 0.08 µg/L and 0.66 ng/L with a linear range (IC20 - IC80) of 0.0028-2.25 µg/L for SAL. The IC50 and LOD of the TRFIA improved approximately 5-fold and 31-fold, respectively, when compared with our previously reported ELISA data. When compared to most other conventional methods, the TRFIA also showed an excellent sensitivity and accuracy for the detection of SAL. Recoveries from 83.4 to 111.3% and standard deviations (RSDs) from 3.9 to 14.0% were observed in various environmental SAL-spiked samples, including river water, paddy water, livestock wastewater, vegetable field soil and rice paddy soil. In addition, the developed TRFIA showed largely consistent with analytic results from UPLC-MS/MS (R2 = 0.9825, n = 15). These results suggest that the proposed TRFIA can be applied as a sensitive and reliable monitoring method to detect trace SAL in environmental samples.

Albuterol Delivery Efficiency in a Pediatric Model of Noninvasive Ventilation With a Single-Limb Circuit.

BACKGROUND: Pediatric patients treated with noninvasive ventilation (NIV) are frequently given aerosol therapy. Limited pediatric data are available on the efficiency of aerosol delivery efficiency. We evaluated the effect of different nebulizers, positions in the single-limb ventilator circuit, and ventilator settings on the efficiency of aerosol delivery in a model of pediatric NIV. We hypothesized that using a vibrating mesh nebulizer, placing the nebulizer after the circuit leak, and not using the highest inspiratory positive airway pressure would increase aerosol delivery efficiency. METHODS: We connected a breathing simulator in series to a low-dead-space filter holder (lung dose) and to an anatomically correct face/airway model of a 5-y-old child. A mask with an entrainment elbow was connected to a ventilator operated in a NIV bi-level mode and assembled with a single-limb heated-wired circuit. Inspiratory/expiratory pressures of either 15/5 or 20/5 cm H2O were used. We studied 3 different jet nebulizers and 2 vibrating mesh nebulizers loaded with albuterol solution (2.5 mg/3 mL). Albuterol was measured with spectrophotometry. The outcome measure was the efficiency of aerosol delivery (ie, lung dose expressed as percentage of the nominal dose). RESULTS: Vibrating mesh nebulizers placed after the exhalation port of the circuit had the highest delivery efficiency, even compared with a vibrating mesh nebulizer integrated into the mask. Placing the nebulizer after the exhalation port of the circuit increased efficiency for all nebulizers. Vibrating mesh nebulizers were more efficient than jet nebulizers, regardless of their position in the circuit. Increasing the inspiratory pressure resulted in a variable effect on aerosol-delivery efficiency. CONCLUSIONS: In a model of pediatric NIV using a single-limb circuit, aerosol delivery devices were more efficient when placed after the exhalation port of the ventilator circuit. Vibrating mesh nebulizers were more efficient than jet nebulizers.

Investigation of deep eutectic solvents as additives to ß-CD for enantiomeric separations of Zopiclone, Salbutamol, and Amlodipine by CE.

The enantioseparation of chiral drugs via CE was first investigated using ß-CD as chiral additive and deep eutectic solvents (DESs) as auxiliary additive. The results showed that improved separation of tested chiral drugs was obtained in the presence of DESs and ß-CD compared to the single ß-CD separation system. With the optimized condition, resolutions of DESs applied ß-CD separation system for rac-Zopiclone, rac-Salbutamol, and rac-Amlodipine increased 3-4.2 times as single ß-CD separation system. The resolutions reached 4.74, 6.37, and 9.67, respectively. The results demonstrate that DESs are viable additives to CD system in CE for the separation of the chiral drugs.

Preparation of salbutamol imprinted magnetic nanoparticles via boronate affinity oriented surface imprinting for the selective analysis of trace salbutamol residues.

Salbutamol (SAL) is one of the most widely abused feed additives in animal husbandry. Selective enrichment of SAL from complex biological samples is a key step for the analysis of SAL. Herein, we present SAL-imprinted magnetic nanoparticles (MNPs) as an ideal sorbent for selective enrichment of SAL. The SAL-imprinted MNPs were prepared according to the newly reported boronate affinity oriented surface imprinting. The prepared imprinted MNPs exhibited several significant advantages, such as excellent selectivity towards SAL, superb tolerance to interferences, fast binding equilibrium and easy manipulation, which made the obtained imprinted MNPs an ideal sorbent for selective enrichment. The feasibility of SAL-imprinted MNPs for real samples was well demonstrated by the analysis of trace SAL in pig liver. Because of their highly desirable properties, the SAL-imprinted MNPs could find more applications in the analysis of SAL.

The use of hair as a long-term indicator of low-dose ß2 agonist treatments in cattle: Implications for growth-promoting purposes monitoring.

The objective of this study was to assess the feasibility of using hair as a long-term indicator of cocktail (low-dose ß2 agonists) treatments in cattle. Six male Simmental cattle were treated with a mixture of low-dose clenbuterol, ractopamine, and salbutamol at dosages of 5.3, 223.3, and 50.0 µg/kg, respectively. The trial lasted for 112 days and included 28 days of treatment and 84 days of withdrawal. Plasma and urine samples taken during the treatment period contained the highest residues, with maximum concentrations of clenbuterol, ractopamine, and salbutamol in plasma of 1.49 ng/mL (Day 21), 43.78 (Day 14) ng/mL, and 8.07 ng/mL (Day 7), respectively, and in urine of 62.40 ng/mL (Day 28), 3995.77 ng/mL (Day 28), and 503.72 ng/mL (Day 1), respectively. On day 42 of withdrawal, the residues of all three ß2 agonists in plasma were below the limit of quantification (LOQ; 0.3 ng/mL for clenbuterol, and 0.5 ng/mL for ractopamine and salbutamol), and in urine samples were below or near the LOQ (the highest being ractopamine at 1.10 ng/mL). The highest concentrations of clenbuterol, ractopamine, and salbutamol in hair were 88.36, 1351.92, and 100.58 ng/g, respectively, on day 14 of withdrawal; and the residues were long-lasting, with 7.64, 28.55, and 8.77 ng/g, respectively, on day 84 of withdrawal. The results of this study demonstrate that hair could be utilized as a long-term indicator of the use of a combination of low-dose ß2 agonists in cattle, which could have implications for growth-promoting purposes monitoring.

The Effect of Different Closed Suction Catheter Designs and pMDI Adapters on Aerosol Delivery in Simulated Adult Mechanical Ventilation With and Without Exhaled Humidity.

BACKGROUND: Closed suction catheters are commonly used with pressurized metered-dose inhalers (pMDIs) in ventilator-dependent patients receiving bronchodilator therapy. Although heated/humidified circuits result in a decrease in drug delivery, no studies have investigated the impact of a closed suction catheter with a pMDI on aerosol delivery during mechanical ventilation. We sought to quantify aerosol delivery with a variety of closed suction catheters and pMDI adapters in a simulated ventilator-dependent adult lung model with and without exhaled humidity. METHODS: A ventilator (tidal volume 450 mL, PEEP 5 cm H2O, breathing frequency 15 breaths/min, peak flow 60 L/min, and bias flow 2 L/min) was attached to 3 closed suction catheter designs (T-piece, double swivel elbow, and a multi-access port) with pMDI adapters (small- and large-unidirectional adapters, a bidirectional adapter, and a built-in port) connected to an endotracheal tube (ETT) with a collecting filter and a passive test lung. To simulate exhaled humidity, a heated humidifier was placed between the collection filter and the distal tip of the ETT, set at 36°C. Experiments without exhaled humidity were performed with no heat and humidification. For each test, 4 puffs of albuterol sulfate (432 µg) were administered. The drug was eluted from the collecting filter and analyzed via spectrophotometer (276 nm). Each experiment was run 3 times. RESULTS: Delivery efficiency of the bidirectional adapter was greatest compared to other adapters tested in this study (P = .01). Aerosol deposition obtained from the combination of the T-piece closed suction catheter and the bidirectional adapter was more than that from the multi-access port (P = .42) and double swivel closed suction catheter (P = .27). Drug delivery without exhaled humidity exceeded values with exhaled humidity by 20-90% using the small unidirectional, large unidirectional, and bidirectional adapters (P = .01, P = .02 and P = .02, respectively). CONCLUSIONS: Closed suction catheter, pMDI spacer/adapters, and exhalation conditions have an impact on drug delivery during simulated adult mechanical ventilation. Aerosol delivery without exhaled humidity appears to report unrealistically high values in comparison to the more accurate values with simulated exhalation humidity.

Determination of ractopamine and salbutamol in pig hair by liquid chromatography tandem mass spectrometry.

A liquid chromatography tandem mass spectrometric method was developed for the determination of two ß-agonists (ractopamine and salbutamol) in pig hair samples. An isotope of ractopamine-d5 or salbutamol-d6 as an internal standard was used to carry out quantitative analysis. Concentrated sodium hydroxide was used to pretreat hair samples and then purified by the solid phase extraction (SPE) procedure. The extracted solution was evaporated and reconstituted for injection in the instrument with electrospray ionization (ESI) operating in a positive multiple-reaction-monitoring (MRM) mode. Ractopamine and salbutamol separation were performed on C18 analytical column under gradient condition. The internal standard calibration curve was linear in the range of concentration from 0.5 to 100 ng mL-1 (R2 > 0.995). Recoveries of this method estimated at three spiked concentrations of 100, 250 and 500 ng mL-1 in pig hair samples, were 79-82% for ractopamine and 77-96% for salbutamol. The corresponding inter-day and intra-day precisions expressed as relative standard deviation (RSD %) were 3.8-6.4% and 3.8-8.6%, respectively. The analytical time for one sample was 8 min. The detection limit of this method was 0.6 and 8.3 ng mL-1 for ractopamine and salbutamol, respectively. This developed method can be applied for monitoring the use of the ß-agonists salbutamol and ractopamine in swine feed incurred pig hair.

Highly efficient detection of salbutamol in environmental water samples by an enzyme immunoassay.

Salbutamol (SAL) has potential hazardous effects on health, and has been found in natural water globally. To protect consumers from risks related to SAL residues, a fast and highly sensitive enzyme-linked immunosorbent assay (ELISA) was developed for the detection of SAL in environmental water samples. SAL hapten was synthesized and conjugated to proteins to be used as the immunogen for producing polyclonal antibodies. Under optimized conditions, the ELISA showed a 50% inhibitory concentration (IC50 value) of 0.466µg/L, with a limit of detection (LOD, IC10 value) of 0.021µg/L. The LOD of the ELISA was 5-fold lower than the maximum residue limits. The average recoveries of SAL from environmental samples, including tap water, river water and wastewater, ranged from 80.1 to 115.6%, which is well within the requirements of residue detection. Moreover, the results of the ELISA correlated well with high-performance liquid chromatography results, with a correlation coefficient of 0.988. These results suggest that the proposed ELISA method has potential application for the monitoring of SAL residues in environmental water samples.

A hybrid molecularly imprinted polymer coated quantum dot nanocomposite optosensor for highly sensitive and selective determination of salbutamol in animal feeds and meat samples.

A hybrid molecularly imprinted polymer (MIP)-coated quantum dot (QD) nanocomposite was synthesized and applied as a fluorescence probe for the highly sensitive and selective determination of salbutamol. The hybrid MIP-coated QD nanocomposite was synthesized via a copolymerization process in the presence of thioglycolic acid capped CdTe QDs with salbutamol as a template, 3-aminopropyltriethoxysilane as the functional monomer, and tetraethyl orthosilicate as a cross-linker. The optimum molar ratio of template, monomer, and cross-linker was 1:6:20. The fluorescence intensity of the hybrid MIP-coated QDs was efficiently quenched after salbutamol rebound to the recognition sites, as a result of charge transfer from QDs to salbutamol. The synthesized hybrid MIP-coated QD nanocomposite showed high sensitivity and good selectivity toward salbutamol. Under the optimal recognition conditions, the fluorescence intensity was quenched linearly with increasing concentration of salbutamol in the range from 0.10 to 25.0 µg L-1, with a detection limit of 0.034 µg L-1. The hybrid optosensor developed was successfully applied in the determination of salbutamol in animal feeds and meat samples. Satisfactory recoveries were obtained in the range from 85% to 98%, with a standard deviation of less than 8%. Furthermore, the accuracy of the hybrid MIP-coated QD nanocomposite was investigated by comparison with a conventional high-performance liquid chromatography method, with the results obtained with two methods agreeing well with each other. The advantages of this sensing method are simplicity, rapidity, cost-effectiveness, high sensitivity, and good selectivity. Graphical Abstract The synthesis of hybrid MIP-coated QDs nanocomposite.

Discriminating Ability of Abbreviated Impactor Measurement Approach (AIM) to Detect Changes in Mass Median Aerodynamic Diameter (MMAD) of an Albuterol/Salbutamol pMDI Aerosol.

This article reports on results from a two-lab, multiple impactor experiment evaluating the abbreviated impactor measurement (AIM) concept, conducted by the Cascade Impaction Working Group of the International Pharmaceutical Aerosol Consortium on Regulation and Science (IPAC-RS). The goal of this experiment was to expand understanding of the performance of an AIM-type apparatus based on the Andersen eight-stage non-viable cascade impactor (ACI) for the assessment of inhalation aerosols and sprays, compared with the full-resolution version of that impactor described in the pharmacopeial compendia. The experiment was conducted at two centers with a representative commercially available pressurized metered dose inhaler (pMDI) containing albuterol (salbutamol) as active pharmaceutical ingredient (API). Metrics of interest were total mass (TM) emitted from the inhaler, impactor-sized mass (ISM), as well as the ratio of large particle mass (LPM) to small particle mass (SPM). ISM and the LPM/SPM ratio together comprise the efficient data analysis (EDA) metrics. The results of the comparison demonstrated that in this study, the AIM approach had adequate discrimination to detect changes in the mass median aerodynamic diameter (MMAD) of the ACI-sampled aerodynamic particle size distribution (APSD), and therefore could be employed for routine product quality control (QC). As with any test method considered for inclusion in a regulatory filing, the transition from an ACI (used in development) to an appropriate AIM/EDA methodology (used in QC) should be evaluated and supported by data on a product-by-product basis.

A novel approach to study the pMDI plume using an infrared camera and to evaluate the aerodynamic properties after varying the time between actuations.

Plume characteristics, such as temperature and velocity, emitted from pMDIs could significantly affect the dose delivered to the lung. Currently, high speed cameras and thermocouples are used separately to evaluate these parameters. We used a low-noise infrared camera to evaluate both the temperature and velocity of the emitted plume from pMDIs. Additionally, we investigated whether the fine particle fraction (FPF) is affected when time between actuations is varied. We tested three different albuterol sulfate pMDIs: ProAir® HFA, Proventil® HFA, and Ventolin® HFA. The plume and aerodynamic characteristics from these pMDIs were evaluated, after varying the time between actuations (15, 30, 60, and 120s), using the infrared camera and a next generation impactor, respectively. The aerodynamic characteristics were evaluated with and without a valved holding chamber (VHC). ProAir HFA had the softest plume followed by Proventil HFA and Ventolin HFA. Further, Ventolin HFA was slightly cooler and had significantly lower FPF than ProAir HFA and Proventil HFA. All inhalers had higher FPF when used with VHC. Further, we observed that the time between actuations affected the FPF across pMDIs. Moreover, generalized guidelines suggesting one-minute interval between actuations for pMDIs should be reconsidered, with and without a VHC.

Laboratory validation of an LC-MS/MS method for the detection of ractopamine, clenbuterol and salbutamol in bovine and swine muscle at sub-µg kg-1 regulatory limits.

Ractopamine (RAC), is a ß-adrenergic agonist increasingly used in the swine and cattle industry. This compound redirects nutrients to favour leanness rather than fat deposition, improves growth and carcass traits gaining higher economic benefit to producers. Countries around the world are split over whether to allow the use of RAC in meat production. Clenbuterol (CLB) and salbutamol (SLB) are anillinic and phenolic ß-agonists, respectively, with the same capacity of producing economic benefits for the meat sector. However, they are prohibited because of the potentially adverse reactions they can cause in consumers. The three ß-agonist compounds have been included in the Brazilian National Regulatory Survey and consequentially there is an eminent need for reliable methods capable of detecting those substances at the same time and reduce analytical costs. Therefore, an LC-MS/MS method for the simultaneous determination of residual RAC, CLB and SAL in swine and cattle muscle was developed and validated with quantification levels respecting the action levels established for Brazil which are 0.1, 0.2 and 5 µg kg-1 for RAC, CLB and SAL, respectively. Samples were quantified using RAC-d5, CLB-d9 and SLB-d6 as internal standards. The validation was performed according to European Union Decision 2002/657, which includes criteria (CCα, CCß, recovery, repeatability, reproducibility and calibration curve). The method meets the Brazilian regulatory requirement that establishes criteria and procedures for the determination of parameters such as CCα, CCß, precision and recovery. CCα values were 0.02, 0.21 and 5.42 µg kg-1 for RAC, CLB and SAL, respectively, in bovine and swine muscle samples; CCß values were 0.03, 0.22 and 5.8 µg kg-1 for RAC, CLB and SAL, respectively, in bovine and swine muscle samples. Average recoveries fortified with 0.05-7.5 µg kg-1 of the studied ß-agonist leads around 95%. The method was demonstrated to be suitable for the determination of RAC, CLB and SLB in swine and cattle muscle samples.

Effect of the Chain Length of a Modified Layer and Surface Roughness of an Electrode on Impedimetric Immunosensors.

An ultrasensitive label-free impedimetric immunosensor is constructed by modifying a 3-mercaptoproponic acid (MPA) monolayer on highly rough gold nanostructure (AuNS)-electrodeposited screen printed carbon electrodes (SPCEs) for the detection of small molecular weight drugs (SMWDs), such as salbutamol (SAL). The SPCEs preoxidized in a 0.1 M H2SO4 solution (called po-SPCEH2SO4) are electrodeposited with the AuNS to increase the roughness factor to 23.64 ± 1.76, larger than the AuNS/po-SPCENaOH or the AuNS/po-SPCEPBS. Furthermore, the MPA modified layer as a link for the anti-SAL immobilization to give the immunosensors an impedimetric signal-to-noise ratio larger than the 11-mercapto-undecanoic acid- and 16-mercaptohexadecanoic acid-modified layer, due to the lower interfacial impedance of the MPA monolayer. The MPA/AuNS/po-SPCEH2SO4-based immunosensors have a wide linear range of 1 fg mL-1 to 1 ng mL-1 and a limit of detection of 0.6 fg mL-1. Moreover, the immunosensors can practically quantify the SAL concentrations in 1000 times-diluted serum samples with good recovery.