Dispersive liquid-liquid microextraction for the isolation and HPLC-DAD determination of three major capsaicinoids in Capsicum annuum L.

Dispersive liquid-liquid microextraction (DLLME) was combined with high-performance liquid chromatography-diode-array detector (HPLC-DAD) for the extraction and quantitation of three major capsaicinoids (i.e. capsaicin, dihydrocapsaicin and nordihydrocapsaicin) from pepper (Capsicum annuum L.). Chloroform (extraction solvent, 100 μL), acetonitrile (disperser solvent, 1250 μL) and 30 s extraction time were found optimum. The analytes were back-extracted into 300 μL of 50 mM sodium hydroxide/ methanol, 45/55% (v/v), within 15 s before being injected into the instrument. Enrichment factors ranged from 3.3 to 14.7 and limits of detection from 5.0 to 15.0 µg g^-1. Coefficients of determination (R²) and %RSD were higher than 0.9962 and lower than 7.5%, respectively. The proposed method was efficiently applied for the extraction and quantitation of the three capsaicinoids in six cultivars of Capsicum annuum L. with percentage relative recoveries in the range of 92.0%–108.0%. DLLME was also scaled up for the isolation of the three major capsaicinoids providing purity greater than 98.0% as confirmed by liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR) analysis, which significantly reduced the extraction time and organic solvent consumption.     

Cyanogenic Glycoside Analysis in American Elderberry

Cyanogenic glycosides (CNGs) are naturally occurring plant molecules (nitrogenous plant secondary metabolites) which consist of an aglycone and a sugar moiety. Hydrogen cyanide (HCN) is released from these compounds following enzymatic hydrolysis causing potential toxicity issues. The presence of CNGs in American elderberry (AE) fruit, Sambucus nigra (subsp. canadensis), is uncertain. A sensitive, reproducible and robust LC-MS/MS method was developed and optimized for accurate identification and quantification of the intact glycoside. A complimentary picrate paper test method was modified to determine the total cyanogenic potential (TCP). TCP analysis was performed using a camera-phone and UV-Vis spectrophotometry. A method validation was conducted and the developed methods were successfully applied to the assessment of TCP and quantification of intact CNGs in different tissues of AE samples. Results showed no quantifiable trace of CNGs in commercial AE juice. Levels of CNGs found in various fruit tissues of AE cultivars studied ranged from between 0.12 and 6.38 µg/g. In pressed juice samples, the concentration range measured was 0.29–2.36 µg/mL and in seeds the levels were 0.12–2.38 µg/g. TCP was highest in the stems and green berries. Concentration levels in all tissues were generally low and at a level that poses no threat to consumers of fresh and processed AE products.     

Simultaneous Determination of 108 Pesticide Residues in Three Traditional Chinese Medicines Using a Modified QuEChERS Mixed Sample Preparation Method and HPLC-MS/MS

A rapid, efficient, simple, and high-throughput method for the simultaneous determination of 108 pesticide residues in three traditional Chinese medicines (TCMs) was established, comprising an improved QuEChERS method in combination with HPLC-MS/MS based on mixed samples. A quantity of 10 mL of acetonitrile was used as extraction solvent, and 10 mg of amino-modified multi-walled carbon nanotubes (MWCNTs-NH₂) and 150 mg of anhydrous magnesium sulfate (MgSO₄) were selected as sorbents for dispersive solid phase extraction. The performance of the method was verified according to the analytical quality control standards of SANTE/11813/2017 guidelines. With good linearity (R² > 0.9984) in the range of 2–200 μg/L for all pesticides in the selected matrices, and good accuracy, precision, and high sensitivity, the recoveries were in the range of 70–120% for more than 95% of the pesticides, with a relative standard deviation (RSD) of less than 16.82% for all. The limit of detection (LOD) and limit of quantification (LOQ) of the method were 0.01–3.87 μg/kg and 0.07–12.90 μg/kg, respectively, for Fritillaria thunbergii Miq (F. thunbergii), Chrysanthemum Morifolium Ramat (C. morifolium), and Dendrobium officinale Kimura et Migo (D. officinale). The method was successfully applied to 60 batches of actual samples from different regions.     

A Capillary Electrophoresis Method Based on Molecularly Imprinted Solid-Phase Extraction for Selective and Sensitive Detection of Histamine in Foods

In this study, a sensitive capillary electrophoresis (CE) method based on molecularly imprinted solid-phase extraction (MISPE) was proposed to determine histamine in foods. A molecularly imprinted polymer (MIP) synthesized by bulk polymerization was used as the MISPE adsorbent for the selective extraction of histamine. Under the optimal conditions, the MISPE-CE method possessed good linearity for histamine detection in the concentration range of 0.1–100.0 μg/L. The limit of detection and limit of quantification of the method were calculated to be 0.087 μg/L and 0.29 μg/L, respectively. The histamine in spiked rice vinegar and liquor samples were detected by the developed method with recoveries of 92.63–111.00%. The histamine contents in fish, prawn, pork, chicken breast and soy sauce samples were determined using the developed method and a high-performance liquid chromatography method, with no significant difference found between the two methods.     

Development and Comprehensive SPE-UHPLC-MS/MS Analysis Optimization,Comparison, and Evaluation of 2,4-Epibrassinolide in Different Plant Tissues

A determination method for trace 24-epibrassinolide (EBL) in plant tissues was developed using ultra-high-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS). The plant tissue samples were extracted using a methanol–formic acid solution, and the corresponding supernatant was purified with ODS C18 solid-phase extraction column. The extracts were separated using a Zorbax Eclipse Plus C18 (2.1 mm × 50 mm, 1.8 μm) column with methanol and 0.1% formic acid as the mobile phase. The ion source for the mass spectrometry was an electrospray ionization source with positive ion mode detection. The linear range of the target compound was 0.7~104 μg/kg, the limit of detection (LOD) was 0.11~0.37 μg/kg, the limit of quantification (LOQ) was 0.36~1.22 μg/kg, the recovery rate was 84.0~116.3%, and the relative standard deviation (RSD%) was 0.8~10.5. The samples of maize plumule, brassica rapeseed flower, and marigold leaf were detected using the external standard method. The optimization of the extraction method and detection method of EBL improved the detection sensitivity, laid a foundation for the artificial synthesis of EBL, improved the extraction rate of EBL, and provided a theoretical basis for the study of EBL in many plants.     

A Box–Behnken Extraction Design and Hepatoprotective Effect of Isolated Eupalitin-3-O-β-D-Galactopyranoside from Boerhavia diffusa Linn.

The objectives of this study were to optimize and quantify the maximum percentage yield of eupalitin-3-O-β-D-galactopyranosidefrom Boerhavia diffusa leaves using response surface methodology (RSM), as well as to demonstrate the hepatoprotective benefits of the bioactive compound. The Box–Behnken experimental design was utilized to optimize the eupalitin-3-O-β-D-galactopyranoside extraction procedure, which also looked at the extraction duration, temperature, and solvent concentration as independent variables. Boerhaviadiffusa leaves were extracted, and n-hexane, chloroform, ethyl acetate, and water were used to fractionate the dried extracts. The dried ethyl acetate fraction was thoroughly mixed in hot methanol and stored overnight in the refrigerator. The cold methanol was filtered, the solid was separated, and hot methanol was used many times to re-crystallize the solid to obtain pure eupalitin-3-O-β-D-galactopyranoside (0.1578% w/w). The proposed HPTLC method for the validation and quantification of eupalitin-3-O-β-D-galactopyranosidewassuccessfully validated and developed. The linearity (R₂ = 0.994), detection limit (30 ng), and quantification limit (100 ng) of the method, as well as its range (100–5000 ng), inter and intraday precision (0.67% and 0.991% RSD), specificity, and accuracy (99.78% RSD), were all validated as satisfactory. The separation of the eupalitin-3-O-β-D-galactopyranoside band was achieved on an HPTLC plate using toluene:acetone:water (5:15:1 v/v) as a developing system. The Box–Behnken statistical design was used to determine the best optimization method, which was found to be extraction time (90 min), temperature (45 °C), and solvent ratio (80% methanol in water v/v) for eupalitin-3-O-β-D-galactopyranoside. Standard silymarin ranged from 80.2% at 100 µg/mL to 86.94% at 500 µg/mL in terms of significant high hepatoprotection (cell induced with carbon tetrachloride 0.1%), whereas isolated eupalitin-3-O-β-D-galactopyranoside ranged from 62.62% at 500 µg/mL to 70.23% at 1000 µg/mL. More recently, it is a source of structurally unique flavonoid compounds that may offer opportunities for developing novel semi-synthetic molecules.     

Simultaneous Determination of Sulfonamides Antibiotics in Environmental Water and Seafood Samples Using Ultrasonic-Assisted Dispersive Liquid-Liquid Microextraction Coupled with High Performance Liquid Chromatography

The residues and abuse of antibiotics have seriously endangered ecological balance and human health; meanwhile, antibiotics determination is very difficult because of their low levels and multiple categories in complicated matrices. Appropriate sample pretreatment is usually imperative to enrich (ultra)trace antibiotics and eliminate matrix interference prior to chromatographic analysis. Dispersive liquid-liquid microextraction (DLLME) has become an ideal pretreatment technique owing to its simplicity, effectiveness, low-consumption, etc. In this work, an ultrasonic-assisted DLLME (UA-DLLME) was developed for the simultaneous extraction of seven sulfonamides (SAs) antibiotics in environmental water and seafood samples coupled with HPLC-DAD determination. Several parameters affecting UA-DLLME efficiency were systematically optimized, and consequently the SAs were separated and detected within 14.5 min. The obtained limits of detection (LODs) and limits of quantification (LOQs) ranged from 0.7–7.8 μg/L and 2.4–26.0 μg/L for three water samples (seawater, aquaculture wastewater and lake water) and two seafood samples (pomfrets and shrimps). High recoveries (80.0–116.0%) with low relative standard deviations (0.1–8.1%) were achieved for all the tested samples at three spiked levels. Notably, sulfadimethoxine was found at 24.49 μg/L in one seawater sample. The facile, robust and benign DLLME-HPLC method demonstrated promising perspectives for multiresidue analysis of antibiotics.     

Determination of Aniline in Soil by ASE/GC-MS

In this study, a rapid and simple method based on accelerated solvent extraction (ASE) combined with gas chromatography-mass spectrometry (GC-MS) was established to determine the levels of aniline in soil. The matrix spike recovery rates of aniline were investigated by changing several experimental parameters such as vacuum freeze-drying, accelerated solvent extraction, sample transfer, nitrogen-blowing concentration and solvent exchange. Under optimized pretreatment conditions, the linearity of the method ranged from 0.5 to 20 μg mL⁻¹ for aniline, and the correlation coefficient was 0.999. Recoveries of aniline from quartz sand and soil ranged from 76% to 98%, while the precision was excellent with average inter-day and intraday values ranging (n = 6) from 3.1% to 7.5% and 2.0% to 6.9%, respectively. The limits of quantification of the method were 0.04 mg kg⁻¹. Notably, the results show that the method we developed is simple, fast, low cost and can meet the requirements for the determination of aniline in soil samples, sewage sludge, river and pond sediments.     

Chromatographic Determination of Total Selenium in Biofortified Allium sp. following Piazselenol Formation and Micro-Solid-Phase Extraction

Herein, a method based on selective piazselenol formation is applied for total selenium determination in biofortified Allium species. Piazselenol is formed by reacting Se(IV) with an aromatic diamine, namely 4-nitro-1,2-phenylenediamine, in acidic medium. Samples were digested in a nitric acid/hydrogen peroxide open system, followed by selenate reduction in hydrochloric acid. Reaction conditions were optimized in terms of pH, temperature, reaction time, and other auxiliary reagents for interference removal, namely, EDTA and hydroxylamine. For the extraction of the selectively formed 4-nitro-piazselenol, micro-solid-phase extraction (μSPE) was applied, and the analysis and detection of the corresponding complex was performed by HPLC coupled with DAD. An external standard calibration curve was developed (R² = 0.9994) with good sensitivity, and was used to calculate the total selenium content from several Allium plants material, with good intermediate precision (RSD% < 16%). The accuracy of the method was evaluated using both, a comparison with an accepted reference method from our previously published data, as well as three certified reference material with recoveries between 84–126%. The limit of detection was determined to be 0.35 μg/g (in solids) and 1.1 μg/L (in solution), while the limit of quantification was 1.07 μg/g and 3.4 μg/L (in solution). Using the proposed method, selenium content can be quickly and accurately determined in several types of samples. In addition, this study present experimental conditions for overcoming the interferences that might be encountered in selenium determination using piazselenol.     

Ultrasound assisted supramolecular liquid phase microextraction procedure for Sudan I at trace level in environmental samples

A method based on supramolecular liquid phase microextraction has been developed for the preconcentration and determination of trace levels of Sudan I. 1-decanol and tetrahydrofuran were used as supramolecular solvent components. Trace levels of Sudan I were extracted into the extraction solvent phase at pH = 4.0 Analytical parameters such as pH value, supramolecular solvent volume, ultrasonication, centrifugation, model solution volume, matrix effects have been optimized. The limit of detection and the limit of quantification values for Sudan I were calculated as 1.74 μg L⁻¹ and 5.75 μg L⁻¹, respectively. In order to determine the accuracy of the method, addition and recovery studies were carried out to environmental samples.     

A Simple HPLC/DAD Method Validation for the Quantification of Malondialdehyde in Rodent’s Brain

In the present study, a HPLC/DAD method was set up to allow for the determination and quantification of malondialdehyde (MDA) in the brain of rodents (rats). Chromatographic separation was achieved on Supelcosil LC-18 (3 μm) SUPELCO Column 3.3 cm × 4.6 mm and Supelco Column Saver 0.5 μm filter by using a mobile phase acetonitrile (A) and phosphate buffer (20 mM, pH = 6) (B). Isocratic elution was 14% for (A) and 86% for (B). The injection volume (loop mode) was 100 μL with an analysis time of 1.5 min. Flow rate was set at 1 mL/min. The eluted compound was detected at 532 nm by a DAD detector by keeping the column oven at room temperature. The results indicated that the method has good linearity in the range of 0.2–20 μg/g. Both intra- and inter-day precision, expressed as RSD, were ≤15% and the accuracies ranged between ±15%. The lower limit of quantification (LLOQ), stability, and robustness were evaluated and satisfied the validation criteria. The method was successfully applied in a study of chronic toxicology following different treatment regimens with haloperidol and metformin.     

Method Validation and Evaluation of Safrole Persistence in Cowpea Beans Using Headspace Solid-Phase Microextraction and Gas Chromatography

Bioinsecticides are regarded as important alternatives for controlling agricultural pests. However, few studies have determined the persistence of these compounds in stored grains. This study aimed at optimizing and validating a fast and effective method for extraction and quantification of residues of safrole (the main component of Piper hispidinervum essential oil) in cowpea beans. It also sought to assess the persistence of this substance in the grains treated by contact and fumigation. The proposed method used headspace solid-phase microextraction (HS-SPME) and gas chromatography with a flame ionization detector (GC/FID). Factors such as temperature, extraction time and type of fiber were assessed to maximize the performance of the extraction technique. The performance of the method was appraised via the parameters selectivity, linearity, limit of detection (LOD), limit of quantification (LOQ), precision, and accuracy. The LOD and LOQ of safrole were 0.0057 and 0.019 μg kg⁻¹, respectively and the determination coefficient (R²) was >0.99. The relative recovery ranged from 99.26 to 104.85, with a coefficient of variation <15%. The validated method was applied to assess the persistence of safrole residue in grains, where concentrations ranged from 1.095 to 0.052 µg kg⁻¹ (contact) and from 2.16 to 0.12 µg kg ⁻¹ (fumigation). The levels measured up from the fifth day represented less than 1% of the initial concentration, proving that safrole have low persistence in cowpea beans, thus being safe for bioinsecticide use. Thus, this work is relevant not only for the extraction method developed, but also for the possible use of a natural insecticide in pest management in stored grains.     

A QuEChERS-HPLC-MS/MS Method with Matrix Matching Calibration Strategy for Determination of Imidacloprid and Its Metabolites in Procambarus clarkii (Crayfish) Tissues

We developed a method for determination of imidacloprid and its metabolites 5-hydroxy imidacloprid, olefin imidacloprid, imidacloprid urea and 6-chloronicotinic acid in Procambarus clarkii (crayfish) tissues using quick, easy, cheap, effective, rugged, and safe (QuEChERS) and high-performance liquid chromatography-triple quadrupole mass spectrometry. Samples (plasma, cephalothorax, hepatopancrea, gill, intestine, and muscle) were extracted with acetonitrile containing 0.1% acetic acid and cleaned up using a neutral alumina column containing a primary secondary amine. The prepared samples were separated using reverse phase chromatography and scanned in the positive and negative ion multiple reaction-monitoring modes. Under the optimum experimental conditions, spiked recoveries for these compounds in P. clarkii samples ranged from 80.6 to 112.7% with relative standard deviations of 4.2 to 12.6%. The limits of detection were 0.02–0.5 μg·L⁻¹, the limits of quantification were 0.05–2.0 μg·L⁻¹ and the method of quantification was 0.05–2.0 μg·kg⁻¹. The method is rapid, simple, sensitive and suitable for rapid determination and analysis of imidacloprid and its metabolites in P. clarkii tissues.     

A Dilute and Shoot Strategy for Determining Alternaria Toxins in Tomato-Based Samples and in Different Flours Using LC-IDMS Separation

Alternaria toxins are emerging mycotoxins whose regulation and standardization are in progress by the European Commission and the European Committee for Standardization. This paper describes a dilute and shoot approach to determine five Alternaria toxins in selected food samples using liquid chromatography–tandem mass spectrometry (LC-MS/MS). The strategy involves sample extraction with acidified aqueous methanol, followed by a solvent change accomplished via sample evaporation and reconstitution. The quantification is based on isotope dilution, applying all corresponding isotopically labeled internal standards to compensate possible matrix effects of the analysis. The main advantages of the present method over other existing methods includes simple and effective sample preparation, as well as detection with high sensitivity. The five-fold sample dilution can decrease matrix effects, which were evaluated with both external and internal standard methods. The results demonstrated a limit of quantification lower than 1.0 µg/kg for all five analytes for the first time. The newly presented method showed acceptable accuracy (52.7–111%) when analyzing naturally contaminated and spiked standard samples at the described levels. The method was validated for tomato-based and flour samples (wheat, rye, and maize). The absolute recovery ranged from 66.7% to 91.6% (RSD < 10%). The developed method could be an alternative approach for those laboratories that exclude sample cleanup and pre-concentration of state-of-the-art instruments with enhanced sensitivity.     

Optimization of Extraction Conditions and Cytotoxic Activity of Rapanone in Comparison to Its Homologue,Embelin

Rapanone is a plant-derived simple alkyl-dihydroxybenzoquinone structurally close to embelin, a well-known cytotoxic agent. The pharmacological characterization of rapanone is still incomplete, and to fill the data gap, a good source for its acquisition is required to conduct further research. This study aimed to optimize the conditions for the extraction of rapanone from the leaves of white-berried Ardisia crenata Sims. For this purpose, three methods were employed: heat reflux (HRE), shaking (SE), and ultrasound-assisted extraction (UAE), and such parameters as the extraction time, solvent, and the number of extractions from the same sample were set as experimental variables. Furthermore, cytotoxic activity toward prostate cancer, thyroid cancer, and colorectal carcinoma cell lines was investigated and compared with doxorubicin and embelin. The most effective and economical method for the extraction of rapanone was shown to be 20 min UAE with ethyl acetate or chloroform. Rapanone exhibited high cytotoxic activity against PC3 (IC₅₀ = 6.50 μg/mL), Du145 (IC₅₀ = 7.68 μg/mL), FTC133 (IC₅₀ = 6.01 μg/mL), 8505C (IC₅₀ = 7.84 μg/mL), and Caco-2 (IC₅₀ = 8.79 μg/mL) cell lines after 24 h and against the HT29 cell line after 48 h (IC₅₀ = 11.67 μg/mL). Furthermore, it revealed a more favorable safety profile than either its homologue, embelin, or doxorubicin. The set of optimal extraction parameters obtained may be utilized for scientific and industrial purposes to achieve the best rapanone yield. Moreover, this benzoquinone revealed a high cytotoxic activity with good selectivity.     

Determination of Polycyclic Aromatic Hydrocarbons in Traditional Chinese Medicine Raw Material,Extracts, and Health Food Products

The four polycyclic aromatic hydrocarbon markers (PAH4) of benzo[a]anthracene (BaA), chrysene (Chr), benzo[b]fluoranthene (BbF), and benzo[a]pyrene (BaP) are indicators showing polycyclic aromatic hydrocarbon (PAH) contamination levels in Chinese medicine raw materials (CMRMs), extracts and health food products; Samples of herbal medicine, herbal extracts, and food supplements were extracted with n-hexane, then cleaned up sequentially on Florisil and EUPAH solid-phase extraction (SPE) columns. A gas chromatography–mass spectrometry method for the determination of four polycyclic aromatic hydrocarbon markers in Chinese medicine raw material, extracts, and health food products was established; In spiked-recovery experiments, the average recovery was about 78.6–107.6% with a precision of 2.3–10.5%. The limit of quantification (LOQ) and limit of detection (LOD) of the PAH4 markers in this method were 2.0 μg/kg and 0.7 μg/kg, respectively. When the developed method was utilized to determine PAH4 contents in 12 locally available health food products, 3 samples contained over 10.0 μg/kg BaP, and 5 samples contained over 50.0 μg/kg PAH4. The European Union (EU) limits for BaP and PAH4 are 10 and 50.0 μg/kg, respectively; therefore, more attention must be drawn to the exposure risk of BaP and PAH4 in CMRMs, their extracts, and health food products. According to the risk assessment based on the Margin of Exposure (MOE) method, it is recognized that the products mentioned in this study pose a low risk.     

Multiresidue Pesticide Analysis in Tea Using GC–MS/MS to Determine 12 Pesticide Residues (GB 2763-2021)

Pesticides are widely used on tea plants, and pesticide residues are of significant concern to consumers. The National Food Safety Standard Maximum Residue Limits for Pesticides in Food (GB 2763-2021) was recently amended. However, detection methods for pesticides newly added to the list of residues in beverages have not yet been established. For that reason, this study developed a solid-phase extraction (SPE) and gas chromatography–tandem mass spectrometry (GC–MS/MS) method for determining the residues of 12 pesticides, including four newly added, in black and green tea. Sample preparation processes (sample extraction, SPE clean-up, elution solvent, and elution volume) were optimized to monitor these residues reliably. Multiple reaction monitoring (MRM) was used for GC–MS/MS electron impact (EI) mode determination. Finally, satisfactory recoveries (70.7–113.0% for green tea and 72.0–99.1% for black tea) were achieved at three concentrations (10 μg/kg, 20 μg/kg, and 100 μg/kg). The LOQs were 0.04–8.69 μg/kg, and the LODs were 0.01–3.14 μg/kg. This study provides a reliable and sensitive workflow for determining 12 pesticide residues in tea, filling a gap in the newly revised National Standards.     

超声辅助提取-固相萃取土壤中头孢菌素C的高效液相色谱法定量检测

建立了一种采用超声辅助提取(UAE)、强阴离子交换固相萃取(SAX-SPE)净化、高效液相色谱(HPLC)测定土壤中残留头孢菌素C(CPC)简单、快速方法。 样品以超纯水为提取剂,超声辅助提取,SPE柱子以3 mL甲醇和3 mL水活化,采用5 mL的10%甲醇水溶液作为淋洗液,2 mL的5%甲酸水溶液/甲醇溶液(体积比50：50)进行洗脱,高效液相色谱紫外检测器(HPLC-PDA)测定,检测波长λ=254 nm,柱温30 ℃ ,流动相为0.1%甲酸水溶液/甲醇溶液(体积比95：5)对土壤中不同加标浓度的CPC进行检测,方法的回收率在77.9%~98.9%,标准偏差范围为5.0%~6.3%(n=5),方法检出限(LOD)为340.4 μg/kg,定量限(LOQ)为1126.8 μg/kg。 同时采用此方法检测分析新疆某药厂附近阳性土壤样品,不同批次土壤样品结果分别为:检出(低于定量限)、1532.1 μg/kg。     

Caralluma tuberculata N.E.Br Manifests Extraction Medium Reliant Disparity in Phytochemical and Pharmacological Analysis

Solubility of phytoconstituents depends on the polarity of the extraction medium used, which might result in the different pharmacological responses of extracts. In line with this, ethnomedicinally important food plant (i.e., Caralluma tuberculata extracts) have been made in fourteen distinct solvent systems that were then analyzed phytochemically via total phenolic amount estimation, total flavonoid amount estimation, and HPLC detection and quantification of the selected polyphenols. Test extracts were then subjected to a battery of in vitro assays i.e., antioxidants (DDPH scavenging, antioxidant capacity, and reducing power estimation), antimicrobial (antibacterial, antifungal, and antileishmanial), cytotoxic (brine shrimps, THP-1 human leukemia cell lines and normal lymphocytes), and protein kinase inhibition assays. Maximum phenolic and flavonoid contents were computed in distilled water–acetone and acetone extracts (i.e., 16 ± 1 μg/mg extract and 8 ± 0.4/mg extract, respectively). HPLC-DAD quantified rutin (0.58 µg/mg extract) and gallic acid (0.4 µg/mg extract) in methanol–ethyl acetate and methanol extracts, respectively. Water–acetone extract exhibited the highest DPPH scavenging of 36 ± 1%. Total reducing potential of 76.0 ± 1 μg/mg extract was shown by ethanol chloroform while maximum total antioxidant capacity was depicted by the acetone extract (92.21 ± 0.70 μg/mg extract). Maximal antifungal effect against Mucor sp., antileishmanial, brine shrimp cytotoxicity, THP-1 cell line cytotoxicity, and protein kinase inhibitory activities were shown by ethyl acetate-methanol (MIC: 50 µg/disc), n-hexane (IC₅₀: 120.8 ± 3.7 µg/mL), ethyl acetate (LD₅₀: 29.94 ± 1.6 µg/mL), distilled water–acetone (IC₅₀: 118 ± 3.4 µg/mL) and methanol–chloroform (ZOI: 19 ± 1 mm) extracts, respectively. Our findings show the dependency of phytochemicals and bioactivities on the polarity of the extraction solvent and our preliminary screening suggests the C. tuberculata extract formulations to be tested and used in different ailments, however, detailed studies remain necessary for corroboration with our results.     

First Determination of Glycidyl Ester Species in Edible Oils by Reverse-Phase Ultra-Performance Liquid Chromatography Coupled with an Evaporative Light-Scattering Detector

In this work, a new ultra-performance liquid chromatograph-evaporative light-scattering detector (UPLC-ELSD) method for quantitation of glycidyl esters (GE) contents in edible oils is presented. The method features complete separation of five GE species within 20 min by a C18 column and gradient elution with a mobile phase consisting of 85% and 2.5% methanol aqueous solutions. The coefficients of regression (R²) were all ≥0.9999 for the linear-quadratic regression curves of GE species in a concentration range of 5~80 μg/mL. The intraday and interday recoveries (%) of GE species in solvent were in a range of 81.3~107.3%, and the intraday and interday coefficients of variation (CVs, %) were all ≤8.6%. The average recovery (%) of GE species spiked in extra-virgin olive oil samples ranged from 88.3~107.8% and the intermediate precision (CV, %) of ≤14% indicated acceptable accuracy and precision. The method exhibited limit of quantification (LOQ) for each GE species (0.6 μg glycidol equivalents/g oil). The method was applied to determine GE concentrations of six commercial oil samples, and total glycidol equivalents were consistent with data obtained by GC-MS method. This UPLC-ELSD method could be adopted for precursory screening and research purposes to improve food safety when MS detectors are unavailable.