High Solvent Consumption Research Articles

Carbon dioxide expanded liquid: an effective solvent for the extraction of quercetin from South African medicinal plants

BackgroundQuercetin is one of the most important bioflavonoids having positive effects on the biological processes and human health. Typically, it is extracted from plant matrices using conventional methods such as maceration, sonication, infusion, and Soxhlet extraction with high solvent consumption. Our study aimed to optimize the environmentally friendly carbon dioxide-based method for the extraction of quercetin from quince fruit with an emphasis on extraction yield, repeatability, and short extraction time.ResultsA two-step design of experiments was used for the optimization of the key parameters affecting physicochemical properties, including CO2/co-solvent ratio, co-solvent type, temperature, and pressure. Finally, gas expanded liquid combining CO2/ethanol/H2O in a ratio of 10/81/9 (v/v/v) provided the best extraction yield. Extraction temperature 66 °C and pressure 22.3 MPa were the most suitable conditions after careful optimization, although both parameters did not significantly affect the process. It was confirmed by experiments in various pressure and temperature conditions and statistical comparison of obtained data. The optimized extraction procedure at a flow rate of 3 mL/min took 30 min. The repeatability of the extraction method exhibited an RSD of 20.8%.ConclusionsThe optimized procedure enabled very fast extraction in 30 min using environmentally friendly solvents and it was successfully applied to 16 different plant samples, including 14 bulbs and 2 fruits from South Africa. The quercetin content in extracts was quantified using ultra-high performance liquid chromatography (UHPLC) with tandem mass spectrometry. UHPLC hyphenated with high-resolution mass spectrometry was used to confirm chemical identity of quercetin in the analyzed samples. We quantified quercetin in 11 samples of all 16 tested plants. The quercetin was found in Agapanthus praecox from the Amaryllidaceae family and its presence in this specie was reported for the first time.Graphical

Alternative green solvents in sample preparation

Sample treatment is one of the most contaminant steps in the analytical process, mainly due to the high consumption of energy and petrol-based organic solvents, which involve a high environmental impact. One of the most promising strategies to reduce the side effects of the analytical methodology consists of the substitution of organic solvents by a greener alternative. Current green solvents such as subcritical water, supercritical fluids, bio-based solvents, such as methanol, ethanol, d-limonene, and cyrene, surfactant-based solutions including supramolecular solvents and hydrotopes, deep eutectic solvents with special emphasis in natural deep eutectic solvents, and ionic liquids have been evaluated as alternatives to conventional solvents and their advantages and drawbacks have been critically highlighted. Moreover, the different ways employed for evaluate their green character has been discussed including life cycle assessment, CO2 emission or CHEM21 selection guide.

UHPLC coupled with charged aerosol detector for rapid separation of steviol glycosides in commercial sweeteners and extract of Stevia rebaudiana

Natural sweeteners are in high demand as a part of a healthy lifestyle. Among them, sweeteners with decreased caloric value and suitability for diabetes patients are most requested. Extension in their consumption extends the need for their quality control. A fast gradient UHPLC coupled with charged aerosol detection enabling quantitation of stevioside, rebaudioside A-D, and steviolbioside in commercial sweeteners and Stevia rebaudiana plant extracts has been developed. The method was developed to achieve high efficiency, simplicity, versatility, and low solvent consumption. All steviol glycosides were baseline-separated in less than 4 min with a total run time of 7 min. Buffer-free eluents were used in the separations and only 2.45 mL solvent were needed per analysis. The Luna Omega Polar column featuring polar modification of the C18 stationary phase was employed with mobile phases composed of water and acetonitrile for the excellent separation of polar steviol glycosides. The flow rate of the mobile phase 0.35 mL/min, column temperature 50 °C and injection volume 2 µL were used. Critical pair of glycosides, stevioside and rebaudioside A, were baseline separated with a resolution of 2.41. The universal charged aerosol detector allowed quantitation of steviol glycosides with a limit of detection and quantitation 0.15 and 0.5 µg/mL, respectively. Method intra-day precision was less than 2% (RSD), and the recovery was 89.6–105.0% and 93.8–111.4% for plant material and sweetener tablets, respectively. The quantity of steviol glycosides in three out of four commercial sweeteners was 3.0–12.3% higher than declared. The content was about 12.4% less than declared in one sample. But the difference from the labeled content corresponded to trueness and precision of the developed method together with variability of sweeteners production. The most abundant glycoside detected in sweeteners was stevioside followed by rebaudioside A. A leaf-to-stem ratio describing the dominant accumulation of steviol glycosides in leaves affected the differences in the amount of steviol glycosides among plant samples.

Counter-Current Suspension Extraction Process of Lignocellulose in Biorefineries to Reach Low Water Consumption, High Extraction Yields, and Extract Concentrations

The processing of large quantities of water in biorefining processes can lead to immense costs for heating, evaporation, and wastewater disposal. These costs may prohibit the exploitation of alternative products, e.g., xylooligosaccharides from straw, which are regarded as too costly. A new counter-current extractions method is proposed that aims at low solvent (water) consumption, as well as high yields and extract concentrations. This process was evaluated with suspension extraction experiments with steam pretreated wheat straw and the process window analysis based on a mass balance for a washing and a leaching scenario. The latter was conducted with two other suspension extraction processes as a comparison. The equilibration time was found to be well below 10 min. While the suspension extraction with and without recycling need to be designed to achieve a high yield or a high concentration and low solvent consumption, the proposed extraction method can reach all three simultaneously. Thus, this new process is evaluated as a potential method to spare water and downstream costs and allow new processing pathways in second-generation biorefineries.

Development and validation of a reliable LC-MS/MS method for simultaneous determination of deoxynivalenol and T-2 toxin in maize and oats

Herein, a new analytical method for sensitive, rapid and reliable determination of two major trichothecenes: deoxynivalenol (DON) and T-2 toxin (T2) in maize and oats has been successfully established and validated. Acetonitrile/water mixture was used for sample extraction, and two solid phase extraction clean-up routes (i.e., Oasis HLB cartridges and MycoSep 227 columns) were investigated for optimal recoveries and adequate clean-up efficiency. The investigated mycotoxins were separated by ultra-performance liquid chromatography (UPLC) system and quantified by triple quadrupole mass spectrometry (MS/MS) using an electro spray-ionization interface (ESI) operating in positive polarity. MRM experiments were optimized for each trichothecene to generate high sensitive transitions. The method was validated according to European Union Directive: 2002/657/EC. Matrix-matched calibration demonstrated nice linearity in the concentration range of 5–200 µg/kg (T2) and 10–800 µg/kg (DON) with a coefficient of determination (R2) of >0.999. Fortified samples covering the same concentration ranges yielded average recovery values ranged from 85.0 to 95.3% with relative standard deviation (RSD) for the repeatability and reproducibility of <8%. Both mycotoxins exhibited limits of detection (0.04–0.12 µg/kg) and quantification (0.13–0.38 µg/kg) below the maximum limits approved by the European Union (EU). There was no significant ion suppression or enhancement for DON and T2 in both matrices since the matrix effect (ME %) varied between −11.6 to 12.9%. The proposed analytical method was successfully applied to maize and oat samples collected from local Egyptian markets. The key benefit of the current method is its ease of use, avoiding high consumption of solvents being therefore cost-effective, compatible with green analytical chemistry, and time-saving analytical approach.

NMR spectroscopy as a “green analytical method” in metabolomics and proteomics studies

Nuclear magnetic resonance (NMR) is one of two methods (in addition to MS) that predominate in metabolomics and structural analysis. Due to the great demand for this type of research, there is a very high consumption of solvents. Fortunately, most biological studies can be performed using simple protocols that include 10% D2O and 90% H2O. More complex metabolomics studies with polar compound extraction require the use of MeOH or MeCN as reagents for protein precipitation. The most extensive Folch extraction method requires a mixture of solvents (MeOH/H2O/CHCl3) for tissue metabolites and sometimes biofluid extraction. In this review, we show that NMR can be a green method for most biological studies that yields useful information where the overall approach is in line with the GSK (GlaxoSmithKline) solvent sustainability guide.

Green methods for the simultaneous analysis of pharmaceutical mixtures present in disparate concentration ranges: Application to antidiabetic mixtures

Diabetes-mellitus is one of the most common chronic diseases worldwide. Today, many single-pill combinations exist and are indicated especially for diabetic patients who are unable to achieve sufficient glycemic-control with one active component. However, some of these combinations are formulated in concentration ratios that present a real challenge for their analysis and quality-control assessment. For instant, linagliptin (L) and saxagliptin (S) are two oral dipeptidylpeptidase‐4 inhibitors that have recently been formulated with metformin (M) in a ratio of 1:400 with almost no simple analytical methods offered in literature for their analysis in such disparate concentration-ratio. In this study, a simple, fast and green capillary electrophoretic (CE) method was established for the simultaneous determination of L and S with M in pharmaceutical-dosage-forms. The developed method was linear in the range of 2–200 μg.mL−1 for L and S and 10–2000 μg.mL−1 for M with r > 0.9993 with low LOD and LOQ values that allowed the proposed method to be successfully applied for the sensitive determination of S, L and M in their binary-combinations of S-M and L-M in pharmaceutical-dosage-forms despite the large differences in M and L or S doses. Upon comparison with the few reported-methods, the proposed-method offers the advantages of high separation efficiency and low solvent consumption. In addition to using simple aqueous-buffer that adds to the greenness of the method and decrease the cost of operation. The Eco-scale penalty points approach and the Green Analytical Procedure Index were applied to assess the greenness of the developed method. Results revealed that the proposed method is a greener alternative to the reported ones This allows our proposed-method to be used for the routine high-through-put analysis required in quality-control laboratories as an efficient and simple eco- friendly analytical-tool.

Nonlinear behavior in preparative liquid chromatography: A method-development case study for hydroxytyrosol purification.

Preparative liquid chromatography has become an important purification method owing to its advantages of high separation efficiency, good reproducibility, and low solvent consumption. Because overloading in preparative liquid chromatography must be performed to increase the throughput in a cycle, nonlinear chromatographic behavior is observed. Therefore, it is crucial to carefully study nonlinear chromatography for the purification of a given product, which facilitates the efficient optimization of the purification parameters. In this work, a method for the development of a purification method using preparative liquid chromatography based on nonlinear chromatography is proposed. Hydroxytyrosol was selected as the subject for method demonstration. Using methanol and ethanol as organic modifiers, the optimum flow rate was determined on three commercial columns entitled C8 TDE, C18 ME, and C18 TDE, respectively. The curves were fitted with the van Deemter equation, with thorough analysis of the A, B, and C terms. Adsorption isotherms were subsequently studied to explore the distribution of solutes between the stationary and mobile phases at equilibrium. C18 TDE, 5vol% ethanol-water, and 0.2mL/min were selected as the optimal separation material, elution solvent, and flow rate, respectively. Purification of hydroxytyrosol was tentatively confirmed on a C18 TDE column with 1.6% sample loading, 90.98% recovery, and 98.01% purity.

UHPLC Coupled with Charged Aerosol Detector for Rapid Separation of Steviol Glycosides in Commercial Sweeteners and Extract of Stevia Rebaudiana

Natural sweeteners are in high demand as a part of a healthy lifestyle. Among them, sweeteners with decreased caloric value and suitability for diabetes patients are most requested. Extension in their consumption extends the need for their quality control. A fast gradient UHPLC coupled with charged aerosol detection enabling quantitation of stevioside, rebaudioside A-D, and steviolbioside in commercial sweeteners and Stevia rebaudiana plant extracts has been developed. The method was developed to achieve high efficiency, simplicity, versatility, and low solvent consumption. All steviol glycosides were baseline-separated in less than 4 min with a total run time of 7 min. Buffer-free eluents were used in the separations and only 2.45 mL solvent were needed per analysis. The state of the art column Luna Omega Polar featuring polar modification of C18 stationary phase was used for the excellent separation of polar steviol glycosides. Critical pair of glycosides, stevioside and rebaudioside A, were baseline separated with a resolution of 2.41. The universal charged aerosol detector allowed quantitation of steviol glycosides with a limit of detection and quantitation 0.15 and 0.5 µg/mL, respectively. Method intra-day precision was less than 2% (RSD), and the recovery was 89.6-105.0% and 93.8-111.4% for plant material and sweetener tablets, respectively. The quantity of steviol glycosides in three out of four commercial sweeteners was 3.0-12.3% higher than declared. The content was about 12.4% less than declared in one sample. The most abundant glycoside detected in sweeteners was stevioside followed by rebaudioside A. A leaf-to-stem ratio describing the dominant accumulation of steviol glycosides in leaves affected the differences in the amount of steviol glycosides among plant samples.

Development of a High-Throughput Affinity Mass Spectrometry (AMS) Platform Using Laser Diode Thermal Desorption Ionization Coupled to Mass Spectrometry (LDTD-MS).

Affinity selection mass spectrometry (MS) or, simply, affinity mass spectrometry (AMS) is a label-free technology that has been used to identify high-affinity ligands of target proteins of interest by screening against small-molecule compound libraries and identifying molecules that are enriched in the presence of the target protein. We have previously applied Agilent Technology's (Santa Clara, CA) RapidFire solid-phase extraction (SPE)-based high-throughput MS technology to screen small-molecule libraries using AMS. However, SPE-based technologies rely on fluidics for desalting and separation prior to mass analysis with attendant high solvent consumption, relatively high sample volume requirements, risk of sample carryover, and frequent maintenance. To address these challenges, we have established an AMS platform using a laser diode thermal desorption-atmospheric pressure chemical ionization (LDTD-APCI) ionization source (Phytronix, Quebec, Canada) coupled with a SCIEX 5600+ TripleTOF MS (Framingham, MA). We also validated a data-independent acquisition (DIA) Sequential Window Acquisition of All Theoretical Mass Spectra (SWATH-MS) method for the robust detection and analysis of small-molecule affinity hits. An informatics platform developed in-house has resulted in a streamlined data analysis workflow for high-throughput AMS screening campaigns and reduced data processing time without compromising data quality. Finally, 68,000 compounds were screened in a single plate and affinity selected hits were confirmed in an orthogonal enzyme activity assay.

Effective extraction and recovery of linarin from Chrysanthemum indicum L. flower using deep eutectic solvents

Conventional approaches for extraction and enrichment of phytochemicals from plant matrix have many drawbacks such as high organic solvent consumption, low extraction yields, time-consuming and large labor for the enrichment process. In the present study, ultrasound-assisted deep eutectic solvents extraction (UAE-DESs) was developed for extraction and determination of the linarin from Chrysanthemum indicum L. flower (CIF). And a feasible strategy was performed to separate and recover the linarin from denatured DESs using AB-8 macroporous resins as adsorbents. Under the optimal extraction conditions, the maximum linarin extraction yield reached 14.23 mg/g with liquid–solid ratio 32 mL/g, extraction power 340 w, extraction time 32 min and deep eutectic solvent of choline chloride-ethylene glycol with the molar ratio 1:2 and water content 30% (v/v) as a extraction system, which was 1.21 times higher than 80% ethanol extraction. After macroporous resins enrichment, the recovery efficiency of linarin reached 81.55%. In addition, the DESs could be regenerated for extraction process, and macroporous resins could be further used for enrichment process with desired performance. This study provided an effective and convenient extraction and recovery method for linarin determination from CIF which was also applicable to other natural plants.

Ultrasonic-Assisted Extraction of Phalerin from Phaleria macrocarpa: Response Surface Methodology and Artificial Neural Network Modelling

Phaleria macrocarpa is historically treasured remedy for treating various allergies, infections and health complications. Commercial availability of this plant extract, however, remains limited as conventional phytochemical extraction techniques require prolong extraction time, high consumption of solvents, in conjunction to being energy intensive. Herein, this study aimed to statistically optimize the ultrasonic-assisted extraction (UAE) of phalerin from P. macrocarpa using the Box–Behnken design (BBD) and the predictive capability of this approach was compared to a model derived from artificial neural network (ANN). In the optimization experiment, for only three relevant UAE parameters viz. solvent ratio, extraction temperature and solid-to-solvent ratio were examined, for the response of the highest extraction of phalerin. Under an optimized condition (R2 = 0.98) [71% methanol, 1:45 solid-to-solvent ratio (g/mL) and extraction temperature of 47 °C], a satisfactory amount of 4.26 ± 0.51 mg/g of phalerin was attained. Comparison between the RSM and ANN revealed the latter being a better predictive model and yielded an appreciably higher predictive capability (R2 = 0.99) in terms of average absolute deviation, AAD (0.24%) versus RSM (AAD = 1.03%).

A Green HPLC Method for Determination of Nine Sulfonamides in Milk and Beef, and Its Greenness Assessment with Analytical Eco-Scale and Greenness Profile.

Sulfonamides have been widely used in the prevention and clinical treatment of bacterial diseases in livestock and poultry. The use of sulfonamides increases the risk of veterinary drug residues in animal derived foods. The traditional reversed phase liquid chromatography methods for sulfonamides residues detection in animal derived foods have the problem of high consumption of organic solvents. The aim of this study was to establish a green high-performance liquid chromatography method for the detection of sulfonamides residues in different animal-origin foods. The sample extraction solutions were purified by the Agela Cleanert PEP-2 cartridge and analyzed by the high-performance liquid chromatography method using ethanol as the green alternative solvent. The proposed method was validated in terms of linear range (20-1000 μg/kg), limit of detection (3.0-12.3 μg/kg), limit of quantitation (10-43 μg/kg), accuracy (80.7-101.3%), and repeatability and reproducibility (RSD <5.9% and RSD <8.5% respectively). The proposed method is an environmentally friendly, sensitive and reliable high-performance liquid chromatography method for simultaneous determination of sulfonamide residues in animal-origin foods. In this work, we firstly developed a green high-performance liquid chromatography method for simultaneous determination of the residues of nine sulfonamides in milk and beef with ethanol as the green alternative solvent.

Unexpected carbonate salt formation during isolation of an enantiopure intermediate by supercritical fluid chromatography

The enantiomers of a chiral building block to be used in pre-clinical manufacturing were separated using supercritical fluid chromatography (SFC). Despite an extensive evaluation of different columns and solvent combinations followed by a careful optimization of the chromatographic method, the preparative separation suffered from low throughput and high solvent consumption. Consequently, additional improvements were necessary. By utilizing stacked injections, the chromatographic run time was almost halved, and the high solvent consumption was reduced by recycling of the two mobile phase components, carbon dioxide and methanol. The carbon dioxide was reprocessed by the SFC instrument, whereas methanol was evaporated and recycled from the fractions collected. Hence, the originally inefficient separation method was turned into a more sustainable one, and the desired enantiopure intermediate was delivered to be used in the following synthesis of the selected candidate drug. Unfortunately, when the intermediate was used in the subsequent amide coupling, a surprisingly poor yield was obtained. This was caused by an unexpected formation of a stable carbonate salt of the intermediate under the chromatographic conditions used. By removal of the carbonate prior to the amide coupling reaction, the manufacturing campaign could be saved, and the candidate drug was successfully delivered in time.

Sample matrix influence on microdrop hold-up in single drop microextraction

Abstract Single drop microextraction has become a widespread liquid/liquid microextraction technique owing to its simplicity, high preconcentration factor and low consumption of organic solvents in the extraction due to direct introduction of the very low volume of extract into the analytical system. Crucial features ensuring excellent repeatability of single drop microextraction include: solvent volume, solvent type, sample agitation, salts addition, and pH. The influence of sample quality on the microdrop volume and agitation type was studied. Effect of the sample matrix, such as water, acid content samples (orange juice), sample containing alcohol (plum brandy) and protein content sample (milk), on the microdrop hold-up was also investigated. For water analysis, several organic solvents such as chloroform, dichloromethane, tetrachloromethane, tetrachloroethane and chlorobenzene were tested; last three mentioned were suitable for SDME experiments. For milk samples analysis, chlorobenzene microdrop was found to be optimal; advantage of salt addition has been reported. For orange samples, 1:10 dilution was suggested with stable microdrop volumes (toluene) of up to 6 µL for lower stirring rates (100 rpm and 250 rpm). For alcohol-content samples, the change of alcohol percentage of real-life samples had to be considered. A strong influence of the matrix quality on the microdrop stability has been proven.

A simple thin layer chromatography based method for the quantitative analysis of biosurfactant surfactin vis-a-vis the presence of lipid and protein impurities in the processing liquid

Abstract The Bacillus surfactin lipopeptides, widely used as potent biosurfactants, have been increasingly researched for diverse applications, necessitating quantification during the production and purification stages. Lipopeptides are typically quantified by Reversed Phase-High- Performance Liquid Chromatography and High Performance Thin Layer Chromatography but these have inherent limitations of long processing times, high capital investment and high solvent consumption. In this paper, an economical and accessible Thin Layer Chromatography (TLC) method for the analysis of surfactin lipopeptides has been developed, its estimation capability in terms of precision and accuracy thoroughly substantiated, and its robustness under diverse solvent and sampling conditions, as well as its specificity in the presence of multiple bioproducts, comprehensively verified. A linear relationship was observed between surfactin concentration and the area formed by surfactin bands on TLC plates with surfactin concentrations in the range of 0.5 g/L and 3 g/L. The TLC analytical technique was suitable for surfactin analysis when surfactin was dissolved in different solvents and when surfactin was spotted by multiple spotting on TLC plates to accuracy and precision of up to 98% and 95% respectively. Based on inter-day error analysis, reproducible results could be obtained with high accuracy and precision (98% and ≥84% respectively) were obtained when analysis was carried out on different days (two weeks apart). While this technique has been confirmed specifically as an investigative tool for the analysis of surfactin lipopeptides, visualisation of peptides, lipids and the fengycin suggests its further application to the quantification of co-existing impurities and other lipopeptide bioproducts.

Research progress on percolation extraction process of traditional Chinese medicines

Percolation extraction is a conventional extraction method used in the processing of traditional Chinese medicines. After medicinal material powder is placed in a percolation tank, the extraction solvent is continuously added, and percolation extract is collected simultaneously. The percolation equipment is simple. The percolation operation is easy. It is applicable to a wide range of medicinal materials. Components that are unstable under thermal conditions can be effectively extracted. However, there are also disadvantages, such as high solvent consumption, long extraction time, and high energy consumption in subsequent concentration processes. This article mainly reviews the research progress on the common equipment types, affecting factors, parameter optimization methods, and process monitoring. According to analysis on literatures, solvent composition, impregnation time, percolation speed, and solvent consumption are considered as the important factors of percolation processes. At present, near-infrared spectroscopy is widely used in the monitoring of percolation process, and partial least square is a commonly used quantitative modeling method. According to the concept of &quot;Quality by Design&quot;, in-depth investigation of the percolation process mechanism and development of process control methods are future development trends. Therefore, process modeling, process optimization and process monitoring shall be improved. The mechanism models and the empirical models of column chromatography can be used as references to construct the percolation process models. The effect of the quality changes of medicinal materials shall be taken into account when optimizing the percolation process parameters. More simple and easy methods shall be developed to monitor the percolation process status and key properties of percolation extracts.

Prevalence and factors associated to the use of illicit drugs and psychotropic medications among brazilian undergraduates

The aim of this study was to verify the prevalence of use of psychoactive substances (PS) and its associated factors in undergraduate students of a university in southern Brazil. The study was carried out with 830 undergraduate students in the year 2016. The individuals answered a self-administered questionnaire about the PS and its prevalence of daily use, in the last 30 days or at any time of their lives, as well as socioeconomic conditions and academic variables. Caffeine-based energy drinks was the most consumed psychoactive substance (96.3%) among undergraduates in the last 30 days, followed by alcohol (64.0%). Among the illicit drugs most consumed in the last 30 days was marijuana (17.3%), while anxiolytics and amphetamines were the most prevalent psychoactive medicaments in the last 30 days. The prevalence of lifetime illicit drugs used by these students was 41.5%, where we highlight besides marijuana (38.6%) the high consumption of cocaine (7.8%), ecstasy (9.3%) and solvents. Socioeconomic and demographic factors such as gender, have children, religion, and financial background as well as academic variables were associated to recent consumption of these substances. This study concluded there is a high prevalence of use of PS among the undergraduate students, including illicit drugs.

Enrichment of sunflower oil with β‐carotene from carrots: Maximization and thermodynamic parameters of the β‐carotene extraction and oil characterization

The objective of this study was to maximize the ultrasonic-assisted extraction (UAE) of β-carotene for sunflower oil, by adjusting the combinations of the operational variables: temperature, time, and oil to carrot ratio, as well as obtaining extraction kinetics. Enriched sunflower oil and commercial sunflower oil was characterized. A polynomial model was developed to describe the β-carotene content as a function of operating parameters and the results indicated that all variables had an influence on the β-carotene extraction. The maximum values of temperature (55°C), time (60 min), and oil to carrot ratio (30 ml/g) promoted the highest removal of β-carotene. UAE was endothermic, irreversible, and the increase in temperature favored the viability and spontaneity of the process. The oils presented linoleic and oleic acid as the main fatty acids and the enrichment of sunflower oil did not affect the fatty acid profile and the contents of phytosterols and tocopherols. Practical applications The demand for functional foods and the minimization of synthetic additives have stimulated the application of natural active compounds to products, such as β-carotene, which is known for its antioxidant properties. To investigate the extraction of β-carotene, it is important to use efficient technologies that maintain the characteristics of the extract and that overcome the disadvantages of conventional techniques, such as longer extraction time and high solvent consumption. In this context, the enrichment of the sunflower oil with β-carotene from the ultrasound-assisted extraction is highlighted. In addition, optimization studies and thermodynamic analysis contribute to the improvement in the process performance, through the systematic selection of the operational variables, and to the understanding of the behavior of the active compound with the interfaces of the extraction, respectively.

Analytical quality by design development of an ecologically acceptable enantioselective HPLC method for timolol maleate enantiomeric purity testing on ovomucoid chiral stationary phase

Official method in Ph. Eur. for evaluation of timolol enantiomeric purity is normal-phase high performance liquid chromatography (NP-HPLC) method. Compared to other HPLC modes, NP is depicted as quite expensive with high consumption of organic solvents which leads to chronic exposure of analysts to toxic and carcinogenic effects. In order to overcome above-mentioned drawbacks, the aim of this study was to develop new method with better eco-friendly features. This was enabled by using protein type Chiral Stationary Phase (CSP) in reversed-phase mode that required up to 10 % (v/v) of organic solvent. Therefore, an enantioselective HPLC method was developed and validated for quantification of (S)-timolol and its chiral impurity, (R)-isomer. Optimized separation conditions on ovomucoid column were set using Analytical Quality by Design (AQbD) approach in method development. Optimization step was performed following the Box-Behnken experimental plan and the influence of three critical method parameters (CMPs) towards enantioseparation of the above-mentioned peak pair was examined. CMPs included variation of acetonitrile content in the mobile phase (5–10 %, v/v), pH value of the aqueous phase (6.0–7.0) and ammonium chloride concentration in the aqueous part of the mobile phase (10−30 mmol L−1). The most relevant critical method attributes (CMAs) in this case were the separation criterion between studied critical pair and retention factor of the second eluting peak, (S)-timolol. Qualitative Design Space (DS) was defined by Monte Carlo simulations providing adequate assurance of method’s qualitative robustness (π = 95 %). The selected working point situated in the middle of the DS was characterized by following combination of CMPs: acetonitrile content in the mobile phase 7 % (v/v), pH value of the aqueous phase 6.8 and concentration of ammonium chloride in aqueous phase 14 mmol L–1. In the next step, the quantitative robustness was tested by Plackett-Burman experimental design. The validation studies confirmed adequacy of the proposed method for its intended purpose. Finally, Analytical Eco-Scale metric tool was applied to confirm that developed method represents excellent green analytical method compared to the official one.