Search Results (32)

A simple, rapid, and cost-effective method for the determination of benzo[a]pyrene (BaP) in edible oil was developed and validated. Nickel oxide-deposited silica (SiO₂@NiO) was employed as a solid-phase extraction (SPE) adsorbent for the extraction of BaP from edible oil, followed by high-performance liquid chromatography–diode array detector (HPLC-DAD) analysis of BaP. The edible oil was diluted with n-hexane and directly loaded to SiO₂@NiO for SPE. The n-hexane was also used to clean the fat-soluble interference substance in the edible oil, and BaP was selectively captured using SiO₂@NiO through the electron donor–acceptor interaction. The SPE conditions, including the amount of adsorbent, volume of washing solvent, and type and volume of desorption solvent, were optimized. This SiO₂@NiO-based SPE coupled with the HPLC-DAD method demonstrated good linearity within a BaP concentration range of 6–1875 ng/g in edible oils, with a limit of detection of 1.3 ng/g, spiked recovery of 97.4–105.1%, and relative standard deviation (RSD) of <3.0%. The method was applied to the analysis of BaP in 11 real oil samples (soybean oil, olive oil, corn germ oil, flaxseed oil, walnut oil, sunflower kernel oil, peanut oil, unrefined oil, and high-temperature frying oil), and the results show that the unrefined oil and high-temperature frying oil were at risk of BaP exceeding acceptable level. Full article

Pharmaceutical compounds are small, invisible, and biologically powerful molecules that, due to widespread production and consumption, have become part of the environment, causing long-term adverse effects on biota even at low doses. Advances in sensitive and reliable analytical methods have made their detection possible in complex environmental matrices such as wastewater. Given the large number of synthesized pharmaceuticals with various therapeutic purposes, the occurrence of a synergistic effect is to be expected, interfering with their analysis. Therefore, the challenging analysis is often improved through the application of different sample preparation techniques. This paper includes the development of an SPE-HPLC-DAD method for the determination of eleven pharmaceuticals from water samples. To achieve better recoveries for the specified pharmaceutical (sulfamethoxazole) and possibly other components of the mixture, a sulfamethoxazole-imprinted polymer (MIP-SMETOX) was prepared and used in combination with a commercial sorbent (Oasis HLB) for MIP-SPE-HPLC-DAD. After optimization of the extraction conditions, both methods were validated. The LOD was 0.1 to 0.5 µg/L for SPE-HPLC-DAD and 0.1 to 0.25 µg/L for MIP-SPE-HPLC-DAD, depending on the pharmaceuticals. The matrix effect is different (77–196%) for both methods. A decrease in the signal for sulfamethoxazole (77%) was observed with SPE-HPLC-DAD, while MIP-SMETOX as a sorbent is not suitable for procaine (196%), and this is also the highest matrix effect. To extend the data obtained, additional in silico methods were used to gain deeper insights into the nature and strength of the binding interactions. Both methods (with and without MIP) confirmed their purpose by determining various validation performance features, and the final goal of the developed methods was tested using complex wastewater. The MIP-SMETOX produced justified its production, as the MIP-SPE-HPLC-DAD method is generally slightly better than the method using only a commercial sorbent. Full article

Two HPLC-based analytical methods, one with DAD and the other with an FLD detector, were developed and validated for the simultaneous analysis of B₁, B₂, and B₆ vitamins, both in pharmaceutical gummies and in gastric and intestinal fluids (with water or milk or orange juice). For the detection of B₁ by fluorometry, a pre-column oxidation/derivatization process was accomplished in the presence of B₂ and B₆ vitamins. The methods were performed on an Aqua column (250 mm × 4.6 mm, 5 mm) at 40 °C, with isocratic elution (70% NaH₂PO₄ buffer pH 4.95 and 30% methanol) and a flow rate of 0.9 mL/min. Both were validated according to ICH specifications in terms of linearity (R² > 0.999), accuracy (% Mean Recovery 100 ± 3%) and precision (%RSD < 3.23). For the analysis of the samples, a stability study (in diluents, pH and fluids) was conducted, while for their purification two different extraction procedures, a liquid/solid for the gummies (%Recovery > 99.8%) and a Solid Phase Extraction (SPE) for the Gastrointestinal (G.I.) fluids, (%Recovery 100 ± 5%) were developed. Finally, to investigate whether the co-administration of B-complex with water, orange juice or milk plays a significant role in their release from gummies, a three-phase in vitro digestion protocol was applied. The results did not show significant differences with a slight superiority in the release of B₂ and B₆ with water, while B₁ with orange juice. Full article

Green coffee is essential in many tropical economies. Its cultivation often necessitates using pesticides that can leave behind residues harmful to human health. To ensure consumer safety, the European Community has set strict maximum residue limits (ranging from 0.01 to 1.0 mg/kg) for pesticides in green coffee sold within Europe. However, the lack of official testing methods for neonicotinoids (NEOs) is a problem, as laboratories must spend resources and time developing and validating suitable analytical methods. This study developed and validated a method for the simultaneous analysis of seven NEOs frequently used in coffee cultivation: acetamiprid, clothianidin, dinotefuran, imidacloprid, nitenpyram, thiacloprid, and thiamethoxam. The proposed methodology uses Strata^®-X PRO cartridges (solid-phase extraction) to remove interfering compounds present in the food matrix and high-performance liquid chromatography (HPLC), equipped with a diode array detector (DAD), to determine NEOs. The accuracy profile strategy validated the method’s suitability for the intended application. NEO recovery rates above 97%; negligible matrix effects (>93%); the linearity of the quantification method (R² values above 0.99); relative biases and standard deviations below 5% and 6%, respectively; and an expected error rate less than 8% allowed to consider the method reliable for the intended objectives. Because of its low ecological impact and simple execution, this method can be used in routine analyses. Full article

The mixotrophic dinoflagellate Akashiwo sanguinea is known to have acute toxic effects on multiple marine organisms, while the composition and chemical properties of its toxins remain unclear. In this study, we established a method for separation and purification of A. sanguinea toxins using chromatographic techniques. The acetone extract of A. sanguinea exhibited higher hemolytic activity and shorter incubation time compared to methanol and ethyl acetate extracts. Five fractions were obtained by solid-phase extraction (SPE), of which SPE3 (acetone/water ratio 3:2) and SPE4 (acetone/water ratio 4:1) exhibited the highest hemolytic activities and allelopathic effects. Further purification on SPE3 and SPE4 using reverse-phase high-performance liquid chromatography (RP-HPLC) coupled with a diode array detector (DAD) resulted in 11 subfractions, among which Fr4-5 displayed the strongest hemolytic activity. Nearly all active subfractions exhibited higher hemolytic activities incubated under light than those in the dark (p < 0.05), suggesting that A. sanguinea can produce both photosensitive and non-photosensitive toxins, with the former being the primary contributors to its hemolytic activity. Molecular characterization by UV-Vis, FTIR, and HRMS/MS analysis revealed that the structural features of Fr4-5 were highly consistent with porphyrin analogs and could be derived from chlorophyll c-related precursors. These findings highlight that the photosensitive toxins in A. sanguinea may serve dual roles in stress adaptation and ecological competition, potentially contributing to the formation of the blooms. Full article

The stem bark of black locust (Robinia pseudoacacia L.) was extracted, and nine antioxidant compounds (R1–R9) were detected by high-performance thin-layer chromatography combined with the radical scavenging 2,2-diphenyl-1-picrylhydrazyl (DPPH•) assay, multi-detection, and heated electrospray high-resolution mass spectrometry. For structure elucidation, the methanolic crude extract was fractionated by solid-phase extraction, and the compounds were isolated by reversed-phase high-performance liquid chromatography with diode array detection. The structures of isolated compounds were elucidated by nuclear magnetic resonance and attenuated total reflectance Fourier-transform infrared spectroscopy as well as gas chromatography-mass spectrometry to determine the double bond position. 3-O-Caffeoyl oleanolic acid (R1), oleyl (R2), octadecyl (R3), gadoleyl (R4), eicosanyl (R5), (Z)-9-docosenyl (R6), docosyl (R7), tetracosyl (R8), and hexacosanyl (R9) caffeates were identified. While R1 has been reported in R. pseudoacacia stem bark, the known R3, R5, R7, R8, and R9 are described for the first time in this species, and the R2, R4, and R6 are new natural compounds. All nine caffeates demonstrated antioxidant activity. The antioxidant effects of the isolated compounds R1–R8 were quantified by a microplate DPPH• assay, with values ranging from 0.29 to 1.20 mol of caffeic acid equivalents per mole of isolate. Full article

Spilanthol is a major N-alkylamide constituent of Acmella oleracea (L.) R.K. Jansen with diverse pharmacological properties. We recently showed the applicability of NADES (natural deep eutectic solvents) for the green extraction of spilanthol. However, the purification of targets from NADES poses a challenging step due to their non-volatility. A simple green method to retrieve spilanthol with minimal instrumental effort was devised, fractioning NADES (choline chloride/methylurea, choline chloride/1,2-propanediol, choline chloride/citric acid) and dry ethanolic extracts by SPE on C18 material, eluting merely with ethanolic solutions. The relative distribution of spilanthol and organic adulteration in SPE fractions were detected by HPLC-DAD, followed by scale-up, quantification and purity determination in an NMR-based approach. Isocratic elution with 52% ethanol (v/v) proved suitable in all experiments. The three purest 10 mL fractions combined yielded 12.21 mg spilanthol at 71.65% purity from NADES extract ChCl/P (choline chloride/1,2-propanediol, molar ratio 1:2, +20% m/m water). Ethanolic extract samples showed purities ranging from 77.27 to 80.27% in combined raw fractions. For all samples, purity increased by removing non-soluble substances from organic solutions. Pooled NADES extract fractions showed 89.71% in final samples, ethanolic extracts 87.25 to 91.93%. The highest purities of individual fractions per extract were 89.23 to 94.15%. This cheap and simple purification process is promising to acquire spilanthol for research purposes or as a sample preparation step before HPLC on a semi-preparative to preparative scale, as the substance is highly priced and scarcely available on the market. Organic solvents can be reused, and preliminary scale-up possibilities are shown. Full article

Neonicotinoids (NEOs), used as insecticides against aphids, whiteflies, lepidopterans, and beetles, have numerous detrimental impacts on human health, including chronic illnesses, cancer, infertility, and birth anomalies. Monitoring the residues in food products is necessary to guarantee public health and ecological balance. The present work validated a new method to measure seven neonicotinoid insecticides (acetamiprid ACT, clothianidin CLT, dinotefuran DNT, imidacloprid IMD, nitenpyram NTP, thiacloprid TCP, and thiamethoxan THT) in wheat. The analytical procedure was based on simple and fast wheat sample cleanup using solid-phase extraction (SPE) to remove interferents and enrich the NEOs, alongside the NEOs’ separation and quantification by reverse-phase chromatography coupled with a diode array detector (DAD). The validation process was validated using the accuracy profile strategy, a straightforward decision tool based on the measure of the total error (bias plus standard deviation) of the method. Our results proved that, in the future, at least 95% of the results obtained with the proposed method would fall within the ±15% acceptance limits. The test’s cost-effectiveness, rapidity, and simplicity suggest its use for determining the levels of acetamiprid, clothianidin, dinotefuran, imidacloprid, nitenpyram, thiacloprid, and thiamethoxam in routine analyses of wheat. Full article

A high-performance liquid chromatography with diode array detector (HPLC-DAD) analytical method was developed for the simultaneous detection of 17 phenolic compounds, including phenols, chlorophenols, alkylphenols, and nitrophenols, in two types of water matrices: wastewater and surface water. Prior to HPLC-DAD determination, a solid-phase extraction (SPE) procedure was optimized. The proposed method uses multiwavelength analysis, with the optimum detection wavelengths selected as 268 nm, 280 nm, 386 nm, 304 nm, and 316 nm. The highest resolution was achieved using a chromatographic column, Eclipse XDB-C18 (150 × 4.6 mm, 5 μm), which was kept at 20 °C. The mobile phase consisted of a gradient elution program, with mobile phase A being a 0.1% H₃PO₄ aqueous solution and mobile phase B being acetonitrile. The flow rate was set at 0.6 mL/min. The 17 target phenolic compounds were fully separated in less than 27 min. All compounds showed good linear regression, with correlation coefficients higher than 0.999. The method’s quantitation limits ranged from 4.38 to 89.7 ng/L for surface water and 7.83 to 167 ng/L for wastewater. The recovery rates were in the range of 86.2–95.1% for surface water and 79.1–86.3% for wastewater. The SPE-HPLC-DAD method was proven to be fast, sensitive, accurate, and reproducible. The developed method was successfully applied for the analysis of the 17 phenolic compounds in real surface water and wastewater samples, with phenol, 2,4-DNP, and 2,4-DNP being determined at levels greater than the method’s limits of quantitation (LOQs). The proposed analytical method represents an original technical resource for the simultaneous determination of 17 phenolic compounds in environmental water matrices. Full article

In the past few decades, considerable scientific strides have been made in the subject of drug analysis in human biological samples. However, the risk caused by incorrect drug plasma levels in patients still remains an important concern. This review paper attempts to investigate the advances made over the last ten years in common sample preparation techniques (SPT) for biological samples based on solid sorbents, including solid-phase extraction (SPE) and solid-phase micro-extraction (SPME), and in particular in the field of molecularly imprinted polymers (MIPs), including non-stimuli-responsive and stimuli-responsive adsorbents. This class of materials is known as ‘smart adsorbents’, exhibiting tailored responses to various stimuli such as magnetic fields, pH, temperature, and light. Details are provided on how these advanced SPT are changing the landscape of modern drug analysis in their coupling with liquid chromatography-mass spectrometry (LC-MS) analytical techniques, a general term that includes high-performance liquid chromatography (HPLC) and ultra-high performance liquid chromatography (UHPLC), as well as any variation of MS, such as tandem (MS/MS), multiple-stage (MSⁿ), and high-resolution (HRMS) mass spectrometry. Some notes are also provided on coupling with less-performing techniques, such as high-performance liquid chromatography with ultraviolet (HPLC-UV) and diode array detection (HPLC-DAD) detection. Finally, we provide a general review of the difficulties and benefits of the proposed approaches and the future prospects of this research area. Full article

In this research, a molecularly imprinted polymer (MIP) was synthesized by precipitation polymerization using oxazepam (OZ) as a template molecule and was subsequently applied as a selective sorbent for the extraction of diazepam (DZP) and its metabolites in urine samples using an SPE cartridge. OZ, temazepam (TZ), nordiazepam (NZ) and DZP were analyzed in the final extracts by high-performance liquid chromatography with diode array detection (HPLC-DAD). The SPE extraction steps were optimized, and the evaluation of an imprinting factor was carried out. The selectivity of the method for OZ versus structurally related benzodiazepines (BZDs), such as bromazepam (BRZ), tetrazepam (TTZ) and halazepam (HZ), was investigated. Under the optimum conditions, the proposed methodology provided good linearity in the range of 10–1500 ng/mL, with limit of detection values between 13.5 and 21.1 ng/mL and recovery levels for DZP and its metabolites from 89.0 to 93.9% (RSD ≤ 8%) at a concentration level of 1000 ng/mL. The proposed method exhibited good selectivity, precision and accuracy and was applied to the analysis of urine samples from a real case of DZP intake. Full article

To investigate the effects of the application of bentonite, tannins, and their combination in alcoholic fermentation, Malvazija istarska (Vitis vinifera L.) white grape must was treated with 95 g/L of bentonite, 25 g/L of a hydrolysable tannin preparation, while the third treatment received the aforementioned doses of both agents. Control grape must was fermented without bentonite and exogenous tannins. All of the produced wines were additionally fined after fermentation with doses of bentonite needed to achieve complete protein stability. Wines were analyzed both after fermentation and after additional bentonite fining. Standard physicochemical parameters were determined by the OIV methods, and phenols were analyzed by high-performance liquid chromatography with diode-array detection (HPLC-DAD), while the concentrations of free and bound volatile aroma compounds were obtained after solid-phase extraction (SPE) followed by gas chromatography–mass spectrometry (GC-MS). Bentonite and tannins in fermentation generally reduced the total dose of bentonite needed for complete stabilization. Treatments with bentonite slightly decreased the concentration of total dry extract, while tannins preserved total acidity. The negative effect of bentonite on flavonoids was more severe. Tannins in fermentation preserved more hydroxycinnamoyltartaric acids with respect to control wine, and this effect was additionally enhanced by bentonite. Volatile and bound aroma composition was affected by all the treatments, while the addition of tannins resulted in higher concentrations of several important odoriferous esters, such as ethyl hexanoate, ethyl decanoate, and hexyl acetate. Additional fining with bentonite to complete protein stabilization annulled some of the positive effects observed after fermentation. Full article

The wine industry produces large amounts of grape pomace (GP), a waste that needs to be disposed of properly. Bioactive compounds with high added value can be recovered from GP as an interesting strategy to reduce the environmental impact. Here, two different technologies were employed to recover polyphenol compounds from GP: microwave hydrodiffusion and gravity (MHG) and ultrasound-assisted extraction (UAE). The further purification of UAE and MHG extracts was carried out through solid-phase extraction (SPE) to obtain three fractions, F1, F2 and F3. ATR-FTIR analysis confirmed the presence of sugar and polysaccharide components in F1, as well as non-anthocyanin and anthocyanin compounds in F2 and F3, respectively. Also, the chemical profile was determined by HPLC-UV-DAD, identifying the presence of catechin in F2, and malvidin-3-O-glucoside chloride and cyanidin chloride derivative as the main anthocyanin compounds in F3. The fractions and their parental extracts were characterized for total phenolic content (TPC) and scavenger activity by in vitro assays. We found that F2-MHG and F3-MHG contained phenol contents 6.5 and 8.5 times higher than those of the parental non-fractionated extracts. Finally, F3-MHG (100 μg/mL, w/v) was shown to reduce the proliferation of HT-29 cells. Full article

A solid-phase extraction (SPE) process, followed by an HPLC-DAD method, was developed and validated to quantify short-chain fatty acids (SCFAs) and applied to analyze chicken feces samples. This study aimed to report the use of the STRATA^TM-X-A 96 Well Plate SPE cartridge method as the first step in quantifying SCFAs. A stationary reverse-phase Luna Omega C18 column kept at 40 °C was used, with a gradient elution mobile phase (H₂SO₄ and Acetonitrile, 98:2), a flow rate of 1.2 mL/min, and detection at 210 nm. A mixture of acetic, propionic, and butyric acid was used as thestandard. The method showed a linear relationship, with a coefficient of determination of R² = 0.9987, R² = 0.9985, and R² = 0.9966 for acetic, propionic, and butyric acid, respectively. Concerning sensitivity, an LOD and LOQ of 0.14, 0.14, 0.14 mg/mL and 0.44, 0.45, 0.43 mg/mL were obtained for acetic, propionic, and butyric acid, respectively. According to the sample analysis, the accuracy was 76.05, 95.60, and 81.56% for acetic, propionic, and butyric acid, respectively. The developed method is simple, fast, linear, sensitive, accurate, precise, and robust for the quantification of SCFAs. This could serve as an alternative to conventional methodologies for the determination of these critical components in the intestinal health of chicken feces. Full article

Fruit seeds belonging to the pomegranate cultivar “Granata” were subjected to extraction and oily component analysis, with the aim of obtaining information about their composition. The presence of conjugated isomers of linolenic acid (CLNA isomers) in the oily phase extracted from the seeds gives a high added value to this part of the fruit, which is too often considered and treated as waste. The separated seeds were subjected to a classic Soxhlet extraction with n-hexane or extraction with supercritical CO₂, assisted by ethanol. The resulting oils were evaluated by ¹H and ¹³C-NMR and AP-MALDI-MS techniques. Differences in the triacylglycerols composition, with particular regard to punicic acid and other CLNA content, were studied in depth. Results showed the prevalence of punicic acid in the triacylglycerol mixture up to the 75%, with clear preponderance in the extract by supercritical fluids. Consequently, other CLNA isomers are, altogether, two-fold less represented in the supercritical extract than in the Soxhlet one. The two oily residues were subjected to solid phase extraction (SPE) and to HPLC-DAD analysis for the polyphenolic isolation and characterization. In addition to HPLC analysis, which showed different content and composition, DPPH analysis to evaluate the antiradical potential showed that the extract obtained with supercritical CO₂ was much more active. Full article