Solvent Extraction Procedure Research Articles

Rapid Determination of Methamphetamine, Methylenedioxymethamphetamine, Methadone, Ketamine, Cocaine, and New Psychoactive Substances in Urine Samples Using Comprehensive Two-Dimensional Gas Chromatography.

Background/Objectives: This study evaluates the applicability of a comprehensive two-dimensional gas chromatography-flame ionisation detection (GC×GC-FID) approach for the simultaneous determination of 12 underivatised psychoactive drugs, including new psychoactive substances, that comprised of amphetamine, methamphetamine, mephedrone, 3,4-methylenedioxyamphetamine, 3,4-methylenedioxymethamphetamine, α-pyrrolidinovalerophenone, n-ethylpentylone (ephylone), norketamine, ketamine, 3,4-methylenedioxypyrovalerone, methadone, and cocaine. Methods: Separation was effected using a non-polar first dimension (1D) and a polar second dimension (2D) column, demonstrating an improved separation of drug compounds compared to a polar/non-polar column configuration. Interference-free baseline separation of all psychoactive compounds in a urine matrix was achieved within 8 min. The GC×GC-FID method was validated according to the guidelines defined by Standard Practices for Method Validation in Forensic Toxicology. Results: The calibration curves for the 12 psychoactive drugs were well correlated (r2 > 0.99) within the concentration ranges of 50-1500 ng mL-1. Detection limits of 10-20 ng mL-1 were obtained, and good repeatability and reproducibility (CV < 11.4%) were attained for retention times and peak areas. Method recoveries for the small-scale solvent extraction procedure ranged from 96.9 to 114.5%, and bias was between -3.1% and 14.5%. Conclusions: The validated approach was successfully applied for the determination of these illicit compounds in spiked urine samples of different concentrations, highlighting its potential for rapid forensic drug screening.

A-317 Development of a patient-focused volumetric absorptive microsampling strategy to assess urinary amino acid excretion

Abstract Background Biochemical determinations of urinary amino acid excretion patterns can diagnose specific inborn errors of metabolism (IEM). Clinical management of these genetic metabolic patients may require frequent urine amino acid tests. A simplified process for transporting home collected urine specimens would benefit the IEM community. Volumetric absorptive microsampling (VAMS) is an emergent alternative sampling technique for the collection of dried biological specimens. We describe the development and preliminary validation results of a VAMS strategy for the analysis of creatinine and amino acids (N=22) from microsampled dried urine. Methods Urine creatinine and amino acids were respectively measured in a clinical laboratory setting using established assays on a CobasPro (Roche) and a MassTrak ultra-performance liquid chromatography (UPLC) amino acid analysis (AAA) solution (Waters). Two 30 µL VAMS tips from a Mitra device (Neoteryx) were used to sample each random urine specimen. An ultrasonication-based solvent extraction procedure was optimized to extract the amino acids and creatinine from the VAMS tips. Solvent extracts were subsequently dried by centrifugal evaporation. Extract from one tip was re-dissolved in water and tested for creatinine using an enzymatic method. The other extract was re-dissolved in 0.02 N sodium hydroxide and tested according to the MassTrak AAA procedure, which deploys pre-column AccQTag derivatization with reversed-phase UPLC on a C18 column (2.1 × 150 mm; 1.7 µm) and UV detection at 260 nm. Accuracy, within-batch precision and short-term stability of the optimized VAMS protocol were determined. Results Extraction solvent with a 60:40 (v/v) water:methanol mixture provided the highest average amino acid recovery. This extraction solvent produced higher average amino acid recoveries at pH 11 (89±13%), than pH 3 (82±10%) or pH 7 (72±11%). Creatinine measured from the VAMS tips (N=44) had a consistent negative bias of 20% relative to neat urine (R=0.9905, slope=0.802, intercept=0.02). The creatinine corrected urine amino acid results from the Mitra device were corrected for this observed concentration bias. The accuracy of the amino acid results derived from the VAMS tip vs neat urine was 104 ± 14%. The average within-batch precision (CV%) of the amino acids was 9.8% (range=6.7% (cystine) to 17.6% (arginine)) and 6.2% for creatinine. The average 1 week storage stability at ambient temperature was 91% (range=73% (ornithine) to 107% (glycine)) for the amino acids and 107% for creatinine. The devised VAMS strategy was applied to the analysis of microsampled dried urine from a patient with cystinuria (OMIM no. 220100). Marked elevations in arginine, cystine, lysine and ornithine were respectively observed and are consistent with the biochemical diagnosis of the patient. Conclusions Creatinine corrected amino acid results from the devised VAMS strategy exhibit good precision and accuracy when compared to tradition analyses of neat urine. The potential use of microsampled dried urine for the diagnosis and monitoring of selected IEM has been demonstrated. The analytical and clinical performance of this VAMS protocol must be further validated, including the use of microsampled dried urine from home specimen collections.

A high-resolution 13C NMR approach for profiling fatty acid unsaturation in lipid extracts and in live Caenorhabditiselegans

Unsaturated fatty acids (UFA) play a crucial role in central cellular processes in animals, including membrane function, development, and disease. Disruptions in UFA homeostasis can contribute to the onset of metabolic, cardiovascular, and neurodegenerative disorders. Consequently, there is a high demand for analytical techniques to study lipid compositions in live cells and multicellular organisms. Conventional analysis of UFA compositions in cells, tissues, and organisms involves solvent extraction procedures coupled with analytical techniques such as gas chromatography, MS and/or NMR spectroscopy. As a nondestructive and nontargeted technique, NMR spectroscopy is uniquely capable of characterizing the chemical profiling of living cells and multicellular organisms. Here, we use NMR spectroscopy to analyze Caenorhabditis elegans, enabling the determination of their lipid compositions and fatty acid unsaturation levels both in cell-free lipid extracts and in vivo. The NMR spectra of lipid extracts from WT and fat-3 mutant C. elegans strains revealed notable differences due to the absence of Δ-6 fatty acid desaturase activity, including the lack of arachidonic and eicosapentaenoic acyl chains. Uniform 13C-isotope labeling and high-resolution 2D solution-state NMR of live worms confirmed these findings, indicating that the signals originated from fast-tumbling lipid molecules within lipid droplets. Overall, this strategy permits the analysis of lipid storage in intact worms and has enough resolution and sensitivity to identify differences between WT and mutant animals with impaired fatty acid desaturation. Our results establish methodological benchmarks for future investigations of fatty acid regulation in live C. elegans using NMR.

An in-situ versatile screening method for identifying SVOC sources in indoor environments

Indoor semivolatile organic compounds (SVOCs) pose a substantial threat to human health. However, identifying the sources of these emissions has been challenging owing to the scarcity of convenient and practical on-site methodologies. Herein, a novel method for source screening was proposed using aluminum silicate sampling strips to adsorb SVOCs from the surface air of indoor materials. The adsorbed SVOC levels indicate the emission intensity of these materials into indoor environments. Additionally, compact sampling strips can be readily fixed to any vertical surface using a static sticker, facilitating the characterization of various materials in practical settings. Laboratory-simulated experiments demonstrated the capability of the proposed method to differentiate between source and non-source materials within a 10-cm distance in the same space. In practical scenarios, the primary emission sources identified via this method exhibited a consistent correlation with the contents of the corresponding materials obtained from the traditional solvent-extraction method. As the adsorbed SVOCs were directly transferred to a GC–MS through thermal desorption instead of the solvent-extraction procedure, the proposed method demonstrated several-fold improvements in analytical sensitivity and efficiency. Using this versatile screening technique, some emerging and important SVOC species were identified within specific indoor materials. Eliminating these sources has been demonstrated as an effective approach to mitigate SVOC pollution. Overall, the proposed method offers a powerful tool for managing indoor pollutants and safeguarding human health.

Chemical Characterization of Terpene-Based Hydrophobic Eutectic Solvents and Their Application for Pb(II) Complexation during Solvent Extraction Procedure.

Solvents prepared from natural terpenes (menthol and thymol), as H-bond acceptors, and a series of organic acids (chain lengths of 8, 10, and 14 C atoms), as H-bond donors, were characterized and tested as reaction media for liquid-liquid extraction purposes. Due to their high hydrophobicity, they seem to be promising alternatives to conventional (nonpolar and toxic) solvents, since they possess relatively less toxic, less volatile, and consequently, more environmentally friendly characteristics. Assuming that the equilibrium is established between solvent and analyte during a ligandless procedure, it can be concluded that those nonpolar solvents can efficiently extract nonpolar analytes from the aqueous environment. Previous investigations showed a wide range of applications, including their use as solvents in extractions of metal cations, small molecules, and bioactive compounds for food and pharmaceutical applications. In this work, hydrophobic solvents based on natural terpenes, which showed chemical stability and desirable physicochemical and thermal properties, were chosen as potential reaction media in the liquid-liquid extraction (LLE) procedure for Pb(II) removal from aqueous solutions. Low viscosities and high hydrophobicities of prepared solvents were confirmed as desirable properties for their application. Extraction parameters were optimized, and chosen solvents were applied. The results showed satisfactory extraction efficiencies in simple and fast procedures, followed by low solvent consumption. The best results (98%) were obtained by the thymol-based solvent, thymol-decanoic acid (Thy-DecA) 1:1, followed by L-menthol-based solvents: menthol-octanoic acid (Men-OctA) 1:1 with 97% and menthol-decanoic acid (Men-DecA) 1:1 with 94.3% efficiency.

A multicomponent LC-MS/MS method for drugs of abuse testing using volumetric DBS and a clinical evaluation by comparison with urine

BackgroundDrug testing commonly use urine as a specimen and immunoassays for screening. The need for supervised urine collection has led to an interest in alternative specimens and a need for using mass spectrometry methods already for screening. In addition, mass spectrometry methods allow for broad multipanel screening which of great value because of the increased number of substances that needs to be covered has increased over time. One alternative specimen of interest for drugs of abuse testing is dried blood spots (DBS) and this work aimed at developing multipanel screening methods based on selected reaction monitoring liquid chromatography - mass spectrometry for both urine and dried finger blood as specimens. Materials and MethodsThe urine method comprised 37 analytes and utilised salted out liquid/liquid extraction in 96-well format, respectively, and the blood method comprised 35 analytes, a 10 µL volumetric DBS device and a two-step solvent extraction procedure. In both cases stable isotope labelled internal standards were used for almost all analytes. ResultsThe methods were validated according to forensic standard. The lowest reporting limits were generally set at 100 ng/mL for urine and 1 ng/mL for blood and the accuracy and imprecision were within limits of 15 and 20%. The methods were applied in a clinical study on patients receiving methadone maintenance treatment for opioid dependence. Methadone was detected in all urine and DBS samples, for urine sometimes below the commonly applied screening cutoff limit of 300 ng/mL. In 20 out of 99 cases no other drug was detected in any specimen. The most commonly other detected substances were pregabalin, amphetamine, alprazolam, zopiclone and THCCOOH. Findings in urine and DBS generally agreed well but more positives were detected in DBS. ConclusionMultipanel methods using liquid chromatography – mass spectrometry suitable for clinical drug screening were successfully developed for urine and blood collected by finger-pricking and stored as DBS.

A Continuous Extraction Protocol for the Characterisation of a Sustainably Produced Natural Indigo Pigment.

The sustainable industrial production of indigo necessitates a unique extraction process to separate the plant-derived compounds. Calcium compounds are added to encourage hydrolysis of these precursors and to facilitate the isolation of the final form, resulting in an organic-inorganic composite pigment with unspecified characteristics. In this study, we devised a continuous solvent extraction procedure to fractionate the organic indigoid phase within the composite pigment. Overcoming challenges posed by limited solubility in the common organic solvents, this method allows for the analysis of individual fractions, significantly enhancing resolution. Comprehensive characterisation using spectroscopic analysis, thermogravimetry, and UHPLC-MS/MS revealed the potential for quantifying primary components of the natural pigment and distinct differentiation from the synthetic dye. This approach also holds promise for establishing robust manufacturing practices in the industrial production of natural indigo.

Asphaltene-Derived Graphene Quantum Dots for Controllable Coatings on Glass, Fabrics, and Aerogels.

Graphene quantum dots (GQDs) derived from natural asphaltene byproducts can produce controlled hydrophobic or hydrophilic interfaces on glass, fabrics, and aerogels. A set of facile solvent extraction methods were used to isolate and chemically prepare materials with different surface functionalities from a commercially derived asphaltene precursor. The organic-soluble fraction was used to create hydrophobic and water-repellent surfaces on glass and cotton fabrics. The GQD solutions could also penetrate the pores of a silica aerogel, rendering it hydrophobic. Alternatively, by extracting the more polar fraction of the GQDs and oxidizing their surfaces, we also demonstrate strongly hydrophilic coatings. This work shows that naturally abundant GQD-containing materials can produce interfaces with the desired wettability properties through a straightforward tuning of the solvent extraction procedure. Owing to their natural abundance, low toxicity, and strong fluorescence, asphaltene-derived GQDs could thus be applied, in bulk, toward a wide range of tunable surface coatings. This approach, moreover, uses an important large-scale hydrocarbon waste material, thereby offering a sustainable alternative to the disposal of asphaltene wastes.

Gold recovery from electronic wastes using a solvent extraction/selective back-extraction strategy

ABSTRACT The present lab-scale investigation describes a simple and efficient approach to the selective solvent extraction and recovery of gold, and copper as a by-product, from Waste Printed Circuit Boards (WPCBs) of different brands of used computers. The process comprised of three steps; leaching of scraps in aqua regia, solvent extraction process under optimized conditions, and ultimately selective back-extraction. The analysis of leach solution by inductively coupled plasma revealed, in addition to gold (0.14 wt%), copper (40.0 wt%), tin (11.9 wt%), and Ni (2.3 wt%) were the other main metals in the WPCBs. A solvent extraction procedure using trioctylamine, as extractant, dissolved in kerosene was employed for the extraction of gold as its anionic chloride-complexes from the leach liquor. The parameters affecting this process including hydrochloric acid concentration, equilibrium time, extractant concentration, initial gold concentration in the sample solution, and the aqueous/organic volume ratio were optimized by means of the statistical technique response surface methodology (RSM). Under the optimized extraction conditions, 99.6% of gold and 23.4% of copper were transferred into the organic phase, while the extracted percentage of other metal ions were negligible. Selective back-extraction by the solution 0.1 M NaOH resulted in the selective precipitation of copper, while the raffinate contained just gold ions.

Valorization Strategies in CO2 Capture: A New Life for Exhausted Silica-Polyethylenimine.

The search for alternative ways to give a second life to materials paved the way for detailed investigation into three silica-polyethylenimine (Si-PEI) materials for the purpose of CO2 adsorption in carbon capture and storage. A solvent extraction procedure was investigated to recover degraded PEIs and silica, and concomitantly, pyrolysis was evaluated to obtain valuable chemicals such as alkylated pyrazines. An array of thermal (TGA, Py-GC-MS), mechanical (rheology), and spectroscopical (ATR-FTIR, 1H-13C-NMR) methods were applied to PEIs extracted with methanol to determine the relevant physico-chemical features of these polymers when subjected to degradation after use in CO2 capture. Proxies of degradation associated with the plausible formation of urea/carbamate moieties were revealed by Py-GC-MS, NMR, and ATR-FTIR. The yield of alkylpyrazines estimated by Py-GC-MS highlighted the potential of exhausted PEIs as possibly valuable materials in other applications.

Modulating Ligand-Exchange Dynamics on Metal-Organic Polyhedra for Reversible Sorting and Hybridization of Miktoarm Star Polymers.

The precise synthesis of miktoarm star polymers (MSPs) remains one of the great challenges in synthetic chemistry due to the difficulty in locating appropriate structural templates and polymer grafting/growing strategies with high selectivity and efficiency. Herein, ≈2 nm metal-organic polyhedra (MOPs), constructed from the coordination of isophthalic acid (IPA) and Cu2+ , are applied as templates for the precise synthesis of 24-arm MSPs for their unique logarithmic ligand-exchange dynamics. Six different polymers are prepared with IPA as an end group and they further coordinated with Cu2+ to afford the corresponding 24-arm star homo-polymers. MSPs can be obtained by mixing targeted homo-arm star polymers in solutions upon thermal annealing. The compositions of MSPs can be facilely and precisely tuned by the recipe of the star polymer mixtures used. Interestingly, the obtained MSPs can be sorted into homo-arm star polymers through a typical solvent extraction procedure. The hybridization and sorting process can be reversibly conducted through the cycle of thermal annealing and solvent treatment. The complex coordination framework not only opens new avenues for the facile and precise synthesis of MSPs and MOPs with hybrid functionalities, but also provides the capability to design sustainable polymer systems.

Modulating Ligand‐Exchange Dynamics on Metal‐Organic Polyhedra for Reversible Sorting and Hybridization of Miktoarm Star Polymers

AbstractThe precise synthesis of miktoarm star polymers (MSPs) remains one of the great challenges in synthetic chemistry due to the difficulty in locating appropriate structural templates and polymer grafting/growing strategies with high selectivity and efficiency. Herein, ≈2 nm metal‐organic polyhedra (MOPs), constructed from the coordination of isophthalic acid (IPA) and Cu2+, are applied as templates for the precise synthesis of 24‐arm MSPs for their unique logarithmic ligand‐exchange dynamics. Six different polymers are prepared with IPA as an end group and they further coordinated with Cu2+ to afford the corresponding 24‐arm star homo‐polymers. MSPs can be obtained by mixing targeted homo‐arm star polymers in solutions upon thermal annealing. The compositions of MSPs can be facilely and precisely tuned by the recipe of the star polymer mixtures used. Interestingly, the obtained MSPs can be sorted into homo‐arm star polymers through a typical solvent extraction procedure. The hybridization and sorting process can be reversibly conducted through the cycle of thermal annealing and solvent treatment. The complex coordination framework not only opens new avenues for the facile and precise synthesis of MSPs and MOPs with hybrid functionalities, but also provides the capability to design sustainable polymer systems.

Evaluation of the potential migration of acaricides from stamped beeswax to honey simulating beehive conditions: A pilot study

There is an issue concerning the migration of pesticides from stamped beeswax to other bee products, mainly, honey. This implies that honey and other beehive-based products could be contaminated, which would represent a potential risk to consumers and food safety. To determine the degree to which acaricides can be transferred from the stamped beeswax to the honey, a pilot experiment was designed consisting of placing beeswax that contained acaricides (endogenous content, τ-fluvalinate, chlorfenvinphos, and coumaphos; spiked samples, coumaphos) in contact with multifloral honey in an incubator. Temperature and shaking conditions were settled simulating beehive conditions. The isolation of the analytes from beeswax involved the use of a QuEChERS (quick, easy, cheap, effective, rugged & safe) method, and honey was analyzed through a solvent extraction procedure. Acaricides were determined in the resulting extracts by gas chromatography with mass spectrometry detection. Coumaphos was the only acaricide transferred into honey in beeswaxes with endogenous acaricide content; meanwhile, results of the experiments with spiked beeswax samples showed that the initial concentration present in the contaminated beeswax significantly influences the transfer to honey, as it was observed that in the spiked beeswax sheets with high levels of coumaphos, the migration from beeswax to honey arose.

Optimization of Solvent Oil Extraction from Sandbox (Hura crepitans Linn.) Seeds

Due to increasing demand for food, industrial applications and biodiesel production on vegetable oils, optimization of oil extraction processes has become imperative to obtain maximum oil yield from vegetable oil feedstocks. The effect of process factors: moisture content, roasting temperature, and time, extraction temperature, and time on oil yield from sandbox seed by solvent extraction were investigated using a 5 × 5 Central Composite Rotatable (Experimental) Design of Response Surface Methodology. Results obtained were analyzed using Analysis of Variance (ANOVA) and SPSS statistical tool at (p = 0.05). The optimum conditions predicted were validated by experiments. The American Oil Chemists’ Society AOCS 5-04 standard procedure for solvent extraction was used in the experiment. The oil yield from the sandbox seed ranged from 20.9-53.6%, and was increased at the range (6-10%) moisture content, (80-100°C) roasting temperature, (5-15 min) roasting time, (90-100°C) extraction temperature and (90-150 min) extraction time. The optimum oil yield of 53.6% was obtained at the processing conditions of 10.0% moisture content, 95oC roasting temperature, 15 min roasting time, 65oC extraction temperature, and 90 min extraction time. Mathematical models to predict sandbox seed oil yield at varying process conditions were developed with an R2 (0.81). The optimum extractable oil yield of 57.10% was predicted for sandbox seed at processing conditions of 10.25% moisture content, 98.72oC roasting temperature, 17.63 min roasting time, 67.01oC extraction temperature, and 96.6 min extraction time. The study results provide data for equipment and process designs for oil extraction from the sandbox and other oilseeds.

Applied novel functionality in separation procedure from leaching solution of zinc plant residue by using non-aqueous solvent extraction

Traditional solvent extraction (SX) procedures limit metal separation and purification, which consist of the organic and aqueous phases. Because differences in metal ion solvation lead to distinct distribution properties, non-aqueous solvent extraction (NASX) considerably expands the scope of solvent extraction by replacing the aqueous phase with alternate polar solvents. In this study, an experimental design approach used non-aqueous solvent extraction to extract cobalt from zinc plant residue. The aqueous phase comprises ethylene glycol (EG), LiCl and metal ions. In kerosene, D2EHPA, Cyanex272, Cyanex301, and Cyanex302 extractants were used as a less polar organic phase. Various factors were investigated to see how they affected extraction, including solvent type, extractant type and phase ratio, pH, Co(II) concentration, and temperature. The results revealed that at a concentration of 0.05 M, the Cyanex301 extractant could achieve the requisite extraction efficiency in kerosene. The optimal conditions were chosen as the concentration of Cyanex 301 (0.05 M), the concentration of cobalt (833 ppm), the pH (3.5), and the percent of EG (80%). As a result, during the leaching process, these systems are advised for extracting and separating a combination of various metal ions.

The fate of the organic phase beyond third phase formation

Since the reverse micellar aggregation is responsible for an undesirable third phase formation in a solvent extraction procedure, the fate of organic phase before and after third phase formation has been probed by DLS and FTIR spectroscopic techniques to unveil the insights.

Rapid Detection of Accelerants in Fire Debris Using a Field Portable Mid-Infrared Quantum Cascade Laser Based Analyzer

Arson presents a challenging crime scene for fire investigators worldwide. Key to the investigation of suspected arson cases is the analysis of fire debris for the presence of accelerants or ignitable liquids. This study has investigated the application and method development of vapor phase mid-Infrared (mid-IR) spectroscopy using a field portable quantum cascade laser (QCL) based system for the detection and identification of accelerant residues such as gasoline, diesel, and ethanol in fire debris. A searchable spectral library of various ignitable fluids and fuel components measured in the vapor phase was constructed that allowed for real-time identification of accelerants present in samples using software developed in-house. Measurement of vapors collected from paper material that had been doused with an accelerant followed by controlled burning and then extinguished with water showed that positive identification could be achieved for gasoline, diesel, and ethanol. This vapor phase mid-IR QCL method is rapid, easy to use, and has the sensitivity and discrimination capability that make it well suited for non-destructive crime scene sample analysis. Sampling and measurement can be performed in minutes with this 7.5 kg instrument. This vibrational spectroscopic method required no time-consuming sample pretreatment or complicated solvent extraction procedure. The results of this initial feasibility study demonstrate that this portable fire debris analyzer would greatly benefit arson investigators performing analysis on-site.

Development and validation of a primary IDMS method for the quantification of 12 sulfonamides in meat by using LC-MS/MS

A new liquid chromatography-tandem mass spectrometry (LC-IDMS) method was developed and validated for a simultaneous multi residue analysis of 12 sulfonamides (SAs) in beef meat. Sulfonamides were isolated from meat with a solvent extraction procedure. Reliable quantitative evaluation was accomplished by using each of the sulfonamides as the internal standart of their own isotopes. Matrix-matched calibration curves were used. Studied performance characteristics were linearity, recovery, precision, detection and quantification limits and robustness. Results were compatible to method performance acceptance criteria according to UME. Recovery results were 82-116%. Measurement uncertainty was calculated from “top down” approach”. Relative measurement uncertainty was between 7-14%.

Dosing Methods to Enable Cell-Based In Vitro Testing of Complex Substances: A Case Study with a PAH Mixture.

Cell-based testing of multi-constituent substances and mixtures for their potential adverse health effects is difficult due to their complex composition and physical-chemical characteristics. Various extraction methods are typically used to enable studies in vitro; however, a limited number of solvents are biocompatible with in vitro studies and the extracts may not fully represent the original test article's composition. While the methods for dosing with "difficult-to-test" substances in aquatic toxicity studies are well defined and widely used, they are largely unsuited for small-volume (100 microliters or less) in vitro studies with mammalian cells. Therefore, we aimed to evaluate suitability of various scaled-down dosing methods for high-throughput in vitro testing by using a mixture of polycyclic aromatic hydrocarbons (PAH). Specifically, we compared passive dosing via silicone micro-O-rings, cell culture media-accommodated fraction, and traditional solvent (dimethyl sulfoxide) extraction procedures. Gas chromatography-tandem mass spectrometry (GC-MS/MS) was used to evaluate kinetics of PAH absorption to micro-O-rings, as well as recovery of PAH and the extent of protein binding in cell culture media with and without cells for each dosing method. Bioavailability of the mixture from different dosing methods was also evaluated by characterizing in vitro cytotoxicity of the PAH mixture using EA.hy926 and HepG2 human cell lines. Of the tested dosing methods, media accommodated fraction (MAF) was determined to be the most appropriate method for cell-based studies of PAH-containing complex substances and mixtures. This conclusion is based on the observation that the highest fraction of the starting materials can be delivered using media accommodated fraction approach into cell culture media and thus enable concentration-response in vitro testing.

Estimation of Nutraceutical Astaxanthin by RP-HPLC Method: Development and Validation

Background: Nutraceuticals have always been considered natural and safe supplements that may prevent disease, may substitute prescription drugs, compensate for a poor diet or promote health. Several nutraceutical products in the market today play roles as antioxidants and probiotics. One such nutraceutical is Astaxanthin, a red-pigmented keto-carotenoid, which offers a plethora of uses and health benefits. Astaxanthin, like many carotenoids, is a lipid-soluble pigment which faces solubility and bioavailability issues owing to its lipophilicity, due to which its estimation is challenging. And the analysis of a simple solvent extract of Astaxanthin would produce many ester-astaxanthin “peaks” and utilize costly organic solvents. Additionally, the literature review indicates the use of very complex mobile phases and tedious solvent extraction procedures involved in the estimation of Astaxanthin. &#x0D; Objective: The manuscript describes the development and validation of a rapid, sensitive and specific Reverse Phase-High Performance Liquid Chromatographic (RP-HPLC) method involving Ultraviolet (UV) detection for the determination and quantification of nutraceutical Astaxanthin in bulk and capsule dosage forms. &#x0D; Methods: Chromatography was carried out on an Inertsil ODS C18 (250 x 4.6 mm), 5µ, using a filtered and degassed mixture of Water: Acetonitrile (30:70% v/v) as the mobile phase at a flow rate of 1 mL /min with isocratic elution method and effluent was monitored at 478 nm. &#x0D; Results: The method was linear in the range of 5 – 15 μg mL-1 of Astaxanthin. The method was validated with respect to precision, accuracy, specificity and robustness. The run time of the analyte was 8.45 min. &#x0D; Conclusion: The developed method is simple, specific, precise, accurate, and robust and shows no interference with diluents which proves the adaptability of the method for routine analysis of the nutraceutical Astaxanthin in bulk and marketed formulations.