Performance of global retention models in the optimisation of the liquid chromatographic separation (II): Complex multi-analyte samples.

BACKGROUND: Enhancing the quality control of medicinal plants is a complex challenge due to their rich variety of chemical compounds present at varying and extreme concentrations. Chromatographic fingerprints, which have become essential for characterising these complex natural materials, require achieving optimal separation conditions to effectively maximise the number of detected peaks. The challenges in optimising fingerprints and other complex multi-analyte samples include the unavailability of standards, the presence of unknown constituents and the substantial workload that would require conventional optimisation methods based on models. RESULTS: This work introduces an interpretive optimisation approach which operates on the premise of predicting chromatograms using global models. Initially, a multi-linear gradient experimental design is sequentially executed to accommodate all peaks in the chromatogram in an adequate time window. Following this, a small set of sample peaks (reference peaks) is selected based on their consistent traceability across all chromatograms in the design. Using this reference dataset, a global model is constructed, initially focused solely on the reference peaks and later extended to encompass all detected peaks in the sample. The aim is to find gradients that maximise resolution while minimising analysis time. These optimised gradients are applied successfully to enhance the separation of medicinal plant extracts, with particular emphasis on peppermint and pennyroyal extracts. SIGNIFICANCE: The proposed optimisation relying on global models can be applied to highly complex samples even in the absence of standards, or in cases where standards are available but their use is impractical due to workload constraints. Moreover, in discerning the most promising gradients for highly complex samples, peak purity has demonstrated superior reliability and competitiveness compared to peak capacity as chromatographic objective function.

Ultrasound-assisted matrix solid-phase extraction based on deep eutectic solvents and zinc oxide: Extraction and determination of six active ingredients in Ligustri Lucidi Fructus.

In this study, we propose a novel strategy utilizing deep eutectic solvents (DESs) as both the extraction solvent and dispersing liquid, with nanometer zinc oxide (ZnO) serving as the adsorbent. This method incorporates ultrasound-assisted matrix solid phase dispersion (UA-MSPD) for the extraction of six active components (salidroside, echinacoside, acteoside, specnuezhenide, nuezhenoside G13, and oleanolic acid) from Ligustri Lucidi Fructus samples. The extracts were then analyzed using high-performance liquid chromatography equipped with a diode array detector. The effects of various parameters such as dispersant dosage, DESs volume, grinding time, ultrasonication duration, and eluent volume on extraction recovery were investigated and optimized using a central composite design under response surface methodology. The optimized conditions yielded detection limits ranging from 0.003 to 0.01 mg/g and relative standard deviations of 8.7% or lower. Extraction recoveries varied between 93% and 98%. The method demonstrated excellent linearity for the analytes (R2 ≥ 0.9997). The simple, green, and efficient DESs/ZnO-UA-MSPD technique proved to be rapid, accurate, and reliable for extracting and analyzing the six active ingredients in Ligustri Lucidi Fructus samples.

Flavonoid Profiles in the Pulp of Different Lemon Cultivars and Their Antioxidant Activity Based on UPLC-Q-TOF-MS.

Previous studies have indicated that there may be differences among the varieties of lemon flavonoids, but the details have not yet been made clear, which limits the comprehensive use of different cultivated lemon varieties. In this study, ultra-performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry (UPLC-Q-TOF-MS) and ultraviolet-visible spectroscopy (UV-Vis) were used to investigate the types and contents of flavonoids in the flesh of the main cultivated variety (Eureka) and five common lemon varieties, as well as their in vitro antioxidant activity. A total of 21 compounds were identified, five of which were common compounds. Among them, Verna, Lisbon, and Bearss each have characteristic components that can serve as potential criteria for variety identification. Each of the six varieties of lemon has strong antioxidant activity. The antioxidant activity of different lemon varieties is related to flavonoids. Therefore, Eureka and the other five varieties of lemon are good natural antioxidants, and the cultivation and industrial production of lemons should consider the needs and selection of suitable varieties.

Potency analysis of twelve cannabinoids in industrial hemp via ultrahigh-performance liquid chromatography-tandem mass spectrometry.

RATIONALE: With an increasing appreciation for the unique pharmacological properties associated with distinct, individual cannabinoids of Cannabis sativa, there is demand for accurate and reliable quantification for a growing number of them. In this study, we developed rapid, sensitive, selective, accurate, and validated liquid chromatography-tandem mass spectrometry for the quantification of cannabinoids. METHODS: Crushed industrial hemp flower and leaf sample was extracted by 95% methanol aqueous, sonicated for 30 min. UPLC-MS/MS analysis using Waters Acquity BEH-C18 column and electrospray ionization(ESI) mass spectrometry detector. RESULTS: The method was validated to demonstrate its reproducibility and precision, linearity, recovery investigation, and investigation of matrix effect. The concentration-response relationship for all analyzed cannabinoids were linear with R2 values >0.99, with intra- and inter-day precision and relative errors below 12%. The recovery and matrix effect were measured as 66.1%-104.1% and 70.42%-110.75%. CONCLUSIONS: This study established a UHPLC-MS/MS method for the simultaneous and rapid quantitative determination of twelve cannabinoids in industrial hemp flowers and leaves in 11 min. The method was used to analyze 43 industrial hemp flower and leaf samples, with the data being statistically analyzed. Based on the statistical analysis of the cannabinoids, hemp from different regions and different varieties were well distinguished by the PLS-DA model, with the main contributing substances being cannabidiol, Δ9-tetrahydrocannabinol, and Δ8-tetrahydrocannabinol.

Modeling the liquid-liquid chromatography separation of cannabinoids from hemp extracts.

The separation of cannabinoids from hemp materials is nowadays one of the most promising industrial applications of liquid-liquid chromatography (LLC). Despite various experimental research efforts to purify cannabinoids, there are currently few works on process modeling. Thus, this study aimed to explore a straightforward approach to model the LLC separation of cannabinoids from two hemp extracts with different compositions. The feed materials were simplified to mixtures of preselected key components (i.e., cannabidiol, tetrahydrocannabinol, cannabigerol, and cannabinol). The elution profiles of cannabinoids were simulated using the equilibrium-cell model with an empirical nonlinear correlation. The model parameters were derived from the elution profiles of single-solute pulse injections. For the validation of the proposed approach, LLC separations with the two hemp extracts were performed in descending mode with the solvent system composed of hexane/methanol/water 10/8/2 (v/v/v). The injected sample concentrations were gradually increased from 5 to 100 mg/mL. The results showed that the approach could describe reasonably well the elution behavior of the cannabinoids, with deviations of only 1-2 min between simulated and experimental elution times. However, to improve the prediction accuracy, the model parameters can be refitted to the elution profiles of 3-4 systematically selected pulse injections with specific hemp extracts.

Pressurized liquid extraction followed by high-performance liquid chromatography for determination of beta-ecdysone extracted from Pfaffia glomerata (Spreng.) Pedersen.

Pfaffia glomerata (Spreng.) Pedersen has among its main bioactive compounds saponins, with the phytoestroid ß-ecdysone as its chemical marker. In this study, pressurized liquid extraction (PLE), a green extraction technique used to obtain bioactive compounds from plants, was employed to extract beta-ecdysone from P. glomerata leaves, stems, and roots. The 22 factorial design was used with the variables temperature (333 K and 353 K) and flow rate (1.5 and 2 mL min-1), pressure (300 Bar), time (60 min), and solvent [ethanol and distilled water (70:30 (v/v)] were kept constant for all parts of the plant. The results of experimental responses demonstrated that the factors temperature and flow rate significantly interfere with the yields of leaf (0.499%), root (0.65%) and stem (0.764%) extracts. The latter presented presents the highest yield compared to the other parts of the plant. HPLC results showed the presence of beta-ecdysone in all parts of the plant with concentrations of ß-ecdysone 86.82, 76.53 and 195.86 mg L-1 to leaf, root and stem, respectively. FT Raman results exhibited typical peaks of beta-ecdysone, such as 3310 cm-1, 1654 cm-1, and 1073 cm-1 for all plant parts. Another interesting result was the presence of the peak at 1460 cm-1 in the PLE root extract can be associated with selenium. This foundational knowledge confirms that the PLE extraction process was efficient in obtaining the chemical marker of Pfaffia glomerata in all plant parts.

Comparison of decarboxylation rates of acidic cannabinoids between secretory cavity contents and air-dried inflorescence extracts in Cannabis sativa cv. 'Cherry Wine'.

Studies with secretory cavity contents and air-dried inflorescence extracts of the CBD-rich hemp strain, Cannabis sativa cv. 'Cherry Wine', were conducted to compare the decarboxylation rates of acidic cannabinoids between two groups. The secretory cavity contents acquired from the capitate-stalked glandular trichomes by glass microcapillaries, and inflorescence samples air-dried for 15 days of storage in darkness at room temperature were analysed by high-pressure liquid chromatography. The ratio of acidic cannabinoids to the total cannabinoids was ranging from 0.5% to 2.4% lower in the air-dried inflorescence samples compared to the secretory cavity samples as follows. In the secretory cavity content, the percentage of acidic cannabinoids to the total cannabinoids was measured as 86.4% cannabidiolic acid (CBDA), 6.5% tetrahydrocannabinolic acid (THCA), 4.3% cannabichromenic acid (CBCA), 1.4% cannabigerolic acid (CBGA), and 0.6% cannabidivarinic acid (CBDVA), respectively. In the air-dried inflorescence, however, the acidic cannabinoids were detected with 84% CBDA, 4.8% THCA, 3.3% CBCA, 0.8% CBGA, and 0.3% Δ9-tetrahydrocannabivarinic acid (Δ9-THCVA), respectively. The ratio of cannabidiol (CBD) to cannabidiolic acid (CBDA) was close to 1:99 (w/w) in secretory cavity contents, however, it was roughly 1:20 (w/w) in the air-dried inflorescence. In addition, Δ9-tetrahydrocannabivarin (Δ9-THCV) and Δ9-tetrahydrocannabivarinic acid (Δ9-THCVA) were only detected in the air-dried inflorescence sample, and the ratio of Δ9-THCV to Δ9-THCVA was about 1:20 (w/w). Besides, cannabidivarinic acid (CBDVA) was only observed in the secretory cavity content.

Determination of Flavonolignan Compositional Ratios in Silybum marianum (Milk Thistle) Extracts Using High-Performance Liquid Chromatography.

Milk thistle is one of the most popular ingredients in the liver protection products market. Silymarin is the main component of milk thistle and contains multiple isomers. There have been few studies focusing on the compositional ratios of silymarin isomers. In this study, we developed an HPLC method for the separation and quantification of silymarin isomers, thereby elucidating their compositional ratios. Through the analysis of more than 40 milk thistle extract products on the market, we found that the ratios, specifically Ratio 1 (the silybin B content to the silybin A content, SBNB/SBNA) and Ratio 2 (the sum of the contents of silybin B and isosilybin B to the sum of the contents of silybin A and isosilybin A, (SBNB + IBNB)/(SBNA + IBNA)), are highly consistent across milk thistle extracts, averaging approximately 1.58 and 1.28, respectively. Furthermore, such ratios were verified in milk thistle seed samples. This study introduces significant findings concerning the stable ratios among silymarin isomers in milk thistle extracts and seeds, thereby offering an innovative approach for quality assurance of milk thistle extracts.

Comprehensive Quality Assessment of Brassica napus L. Seeds via HPTLC, LC-QToF, and Anatomical Investigation.

The Brassicaceae family, commonly referred to as cruciferous plants, is globally cultivated and consumed, with the Brassica genus being particularly renowned for its functional components. These vegetables are rich sources of nutrients and health-promoting phytochemicals, garnering increased attention in recent years. This study presents a comprehensive microscopic, chromatographic, and spectroscopic characterization of Brassica napus L. seeds from Kazakhstan aimed at elucidating their morphological features and chemical composition. Microscopic analysis revealed distinct localization of flavonoids, total lipids, and alkaloids. High-performance thin-layer chromatography (HPTLC) analysis of seed extracts demonstrated a complex chemical profile with significant quantities of non-polar compounds in the hexane extracts. Additionally, methanolic extracts revealed the presence of diverse chemical compounds, including alkaloids, flavonoids, and glucosinolates. The chemical composition exhibited varietal differences across different Brassica species, with B. napus L. seeds showing higher concentrations of bioactive compounds. Furthermore, liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QToF-MS) analysis provided insights into the chemical composition, with sinapine isomers, feruloyl, and sinapoyl choline derivatives as major compounds in the seeds. This study contributes to a better understanding of the chemical diversity and quality control methods' approximations of B. napus L. seeds, highlighting their importance in functional food and nutraceutical applications.

Identifying ligands directly interacting with target protein in medicinal herbs by metabolomic analysis of T2-filtered HSQC spectra.

A protocol for efficiently identifying ligands directly interacting with a target protein in complex extracts of medicinal herbs was proposed by combining an adapted 2D perfect-echo Carr-Purcell-Meiboom-Gill heteronuclear single quantum correlation (PE-CPMG HSQC) spectrum with metabolomic analysis. PE-CPMG HSQC can suppress the signal interference from the target protein, allowing more accurate peak quantification than conventional HSQC. Inspired from untargeted metabolomics, regions of interest (ROIs) are constructed and quantified for the mixture or complex extract samples with and without a target protein, and then a binding index (BI) of each ROI is determined. ROIs or corresponding peaks significantly perturbed by the presence of the target protein (BI ≥1.5) are detected as differential features, and potential binding ligands identified from the differential features can be equated with bioactive markers associated with the 'treatment' of the target protein. Quantifying ROI can inclusively report the ligand bindings to a target protein in fast, intermediate and slow exchange regimes on nuclear magnetic resonance (NMR) time scale. The approach was successfully implemented and identified Angoroside C, Cinnamic acid and Harpagoside from the extract of Scrophularia ningpoensis Hemsl. as ligands binding to peroxisome proliferator-activated receptor γ. The proposed 2D NMR-based approach saves excess steps for sample processing and has a higher chance of detecting the weaker ligands in the complex extracts of medicinal herbs. We expect that this approach can be applied as an alternative to mining the potential ligands binding to a variety of target proteins from traditional Chinese medicines and herbal extracts.

The impact of time and storage conditions on samples containing OC pepper sprays on the quantitative ratios of capsaicinoids.

Pepper sprays of the OC type constitute the majority of self-defense sprays available on the market. The active ingredient in these preparations is pepper extract: Oleoresin Capsicum, which contains capsaicinoids - natural compounds with irritant properties. Preparations from OC pepper sprays can be distinguished based on differences in the quantitative ratios of four main capsaicinoids: capsaicin, dihydrocapsaicin, nordihydrocapsaicin, and nonivamide. This raises the question whether information on the quantitative ratios of capsaicinoids can also provide answers to questions regarding comparisons of traces of OC preparations, such as whether traces revealed on the clothing of the victim could originate from an OC spray secured from the suspect, or whether traces on the clothing of the suspect and the victim could come from the same pepper spray. Such comparisons would be viable only if the capsaicinoid profile remained unchanged during evidence storage and as a result of solvent extraction from the tested material. The aim of the presented research was to determine if this is indeed the case. Model aging experiments were conducted to examine whether the capsaicinoid profile in traces of OC preparations changed over time and whether solvent extraction affected this profile. Samples of five different OC preparations were applied to cotton swabs, which, after the evaporation of volatile solvents, were placed in three types of packaging with varying levels of tightness and transparency (tight amber vials, polyethylene bags, paper envelopes). These prepared samples underwent solvent extraction with methanol and analysis using gas chromatography - mass spectrometry, after 28, 84, 147, 196, 252, and 301 days from preparation. The likelihood ratio (LR) was applied as a statistical tool to investigate the data obtained. The LR model was computed using the three variables based on the relative content of nordihydrocapsaicin, nonivamide, and dihydrocapsaicin. The cotton swabs used in the experiments served as a model for both the swabs used by the police for securing liquid evidence and the cotton clothing of individuals sprayed with OC pepper sprays. The findings of the conducted studies suggest that the quantitative relationships of capsaicinoids indeed change over time, both in preparations stored in original containers and in traces of these preparations present on clothing. For traces of OC preparations secured on swabs or present on clothing, these changes are more significant the longer the sample is stored and the less airtight the packaging used.

Chemical Standardization of Milk Thistle (Silybum marianum L.) Extract Using UHPLC-MS/MS and the Method of Standard Addition.

Extracts prepared from the seeds of the medicinal plant milk thistle [Silybum marianum (L.) Gaertn. (Asteraceae)] are widely used as dietary supplements due to anti-inflammatory, antitumor, and hepatoprotective effects. Called silymarin, the main components of lipophilic extracts of milk thistle seeds are flavonoids and flavonolignans including silybin A, silybin B, isosilybin A, isosilybin B, silydianin, silychristin, taxifolin, and 2,3-dehydrosilybins. The aim of this study was to develop a method based on UHPLC-MS/MS for the chemical authentication and standardization of milk thistle silymarin. Validation included the method of standard addition to account for the lack of a blank matrix. Potential matrix effects were investigated by analyzing silymarin standards dissolved only in the initial UHPLC mobile phase. Measurements of six flavonolignans and taxifolin in the milk thistle extract using UHPLC-MS/MS with standard addition or external standard calibration produced similar results for all analytes except silydianin and 2,3-dehydrosilybin B, which showed significant peak enhancement during negative ion electrospray due to botanical matrix effects. The UHPLC-MS/MS-based method of standard addition requires <10 min per injection and is suitable for the standardization of silymarin from milk thistle in support of preclinical and clinical studies of safety and efficacy.

Validation of an Isocratic HPLC Method for Simultaneous Estimation of Major Phytosterols in Prunus spinosa L. extracts.

This study aimed to develop a rapid method for separation of stigmasterol, campesterol and ß-sitosterol in Prunus spinosa L. (sloe) fruit extracts by High Performance Liquid Chromatography system. Samples were prepared by Soxhlet extraction method and separated on a high strength silica C18 column using acetonitrile-methanol mobile phase and Photodiode Array Detector. The optimized method resulted in a linear calibration curve ranging from 1.7 ng mL-1 to 130 ng mL-1 for all three phytosterols. Analyses of internal and external phytosterol standards showed good linearity (R2 of 0.998 to 0.999); LOD and LOQ were determined to be 2.33×10-7-2.18×10-4 and 7.07×10-7-6.60×10-4 mg mL-1, respectively. Repeatability and reproducibility precision analyses showed acceptable values of RSD %. ß-sitosterol was the predominant phytosterol (51.53-81.03 % of total) among all samples. Method validation parameters indicated that this analytical method can be applied for accurate and precise determination of campesterol, stigmasterol and ß-sitosterol, in selected extracts.

Comprehensive identification strategy for rapid profiling of chemical constituents using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry with Rhubarb as an example.

In this study, the collision induced dissociation tandem mass spectrometry (CID-MS/MS) fragmentation pathway of chemical components in rhubarb was wholly explored using 34 standards by UHPLC-QTOF-MS/MS in negative ion mode. In consequently, the diagnostic product ions for speedy screening and categorization of chemical components in rhubarb were ascertained based on their MS/MS splitting decomposition patterns and intensity analysis. According to these findings, a fresh two-step data mining strategy had set up. The initial key step involves the use of characteristic product ions and neutral loss to screen for different types of substituents and basic skeletons of compounds. The subsequent key step is to screen and classify different types of compounds based on their characteristic product ions. This method can be utilized for rapid research, classification, and identification of compounds in rhubarb. A total of 356 compounds were rapidly identified or tentatively characterized in three rhubarb species extracts, including 150 acylglucoside, 125 anthraquinone, 65 flavanols and 15 other compounds. This study manifests that the analytical strategy is feasible for the analysis of complex natural products in rhubarb.

Ultrafast, Selective, and Highly Sensitive Nonchromatographic Analysis of Fourteen Cannabinoids in Cannabis Extracts, Δ8-Tetrahydrocannabinol Synthetic Mixtures, and Edibles by Cyclic Ion Mobility Spectrometry-Mass Spectrometry.

The diversity of cannabinoid isomers and complexity of Cannabis products pose significant challenges for analytical methodologies. In this study, we developed a method to analyze 14 different cannabinoid isomers in diverse samples within milliseconds by leveraging the unique adduct-forming behavior of silver ions in advanced cyclic ion mobility spectrometry-mass spectrometry. The developed method achieved the separation of isomers from four groups of cannabinoids: Δ3-tetrahydrocannabinol (THC) (1), Δ8-THC (2), Δ9-THC (3), cannabidiol (CBD) (4), Δ8-iso-THC (5), and Δ(4)8-iso-THC (6) (all MW = 314); 9α-hydroxyhexahydrocannabinol (7), 9ß-hydroxyhexahydrocannabinol (8), and 8-hydroxy-iso-THC (9) (all MW = 332); tetrahydrocannabinolic acid (THCA) (10) and cannabidiolic acid (CBDA) (11) (both MW = 358); Δ8-tetrahydrocannabivarin (THCV) (12), Δ8-iso-THCV (13), and Δ9-THCV (14) (all MW = 286). Moreover, experimental and theoretical traveling wave collision cross section values in nitrogen (TWCCSN2) of cannabinoid-Ag(I) species were obtained for the first time with an average error between experimental and theoretical values of 2.6%. Furthermore, a workflow for the identification of cannabinoid isomers in Cannabis and Cannabis-derived samples was established based on three identification steps (m/z and isotope pattern of Ag(I) adducts, TWCCSN2, and MS/MS fragments). Afterward, calibration curves of three major cannabinoids were established with a linear range of 1-250 ng·ml-1 for Δ8-THC (2) (R2 = 0.9999), 0.1-25 ng·ml-1 for Δ9-THC (3) (R2 = 0.9987), and 0.04-10 ng·ml-1 for CBD (4) (R2 = 0.9986) as well as very low limits of detection (0.008-0.2 ng·ml-1). Finally, relative quantification of Δ8-THC (2), Δ9-THC (3), and CBD (4) in eight complex acid-treated CBD mixtures was achieved without chromatographic separation. The results showed good correspondence (R2 = 0.999) with those obtained by gas chromatography-flame ionization detection/mass spectrometry.

UPLC-Q-TOF-MS/MS combined with machine learning methods for screening quality indicators of Hypericum perforatum L.

Hypericum perforatum L. (HPL), also known as St. John's wort, is one of the extensively researched domestically and internationally as a medicinal plant. In this study, non-targeted metabolomics combined with machine learning methods were used to identify reasonable quality indicators for the holistic quality control of HPL. First, the high-resolution MS data from different samples of HPL were collected, and visualized the chemical compounds through the MS molecular network. A total of 122 compounds were identified. Then, the orthogonal partial least squares-discriminant analysis (OPLS-DA) model was established for comparing the differences in metabolite expression between flower, leaf, and branches. A total of 46 differential metabolites were screened out. Subsequently, analyzing the pharmacological activities of these differential metabolites based on protein-protein interaction (PPI) network. A total of 25 compounds associated with 473 gene targets were retrieved. Among them, 13 highly active compounds were selected as potential quality markers, and five compounds were ultimately selected as quality control markers for HPL. Finally, three different classifiers (support vector machine (SVM), random forest (RF), and K-nearest neighbor (KNN)) were used to validate whether the selected quality control markers are qualified. When the feature count is set to 122 and 46, the RF model demonstrates optimal performance. As the number of variables decreases, the performance of the RF model degrades. The KNN model and the SVM model also exhibit a decrease in performance but still manage to satisfy the intended requirements. The strategy can be applied to the quality control of HPL and can provide a reference for the quality control of other herbal medicines.

Buriti (Mauritia flexuosa) shell flour: Nutritional composition, chemical profile, and antioxidant potential as a strategy for valuing waste from native Brazilian fruits.

The Cerrado is one of the most biodiverse biomes in the world, characterized by a wealth of native fruits with unique nutritional characteristics. In this sense, the social, economic, and environmental importance of fully utilizing food is widely recognized. Therefore, generally considered waste, fruit shells can be transformed into a coproduct with high added value. The objective of this work was to carry out a comprehensive assessment of the physicochemical properties, carbohydrate and fatty acid profile, phytochemical compounds, phenolic profile, and antioxidant potential of the recovered extracts of buriti (Mauritia flexuosa) shells in natura and dehydrated at 55 °C (flour). In addition, the functional properties were verified by thermogravimetric analysis (TGA), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR) from buriti shell flour. The results indicated high fiber content and energy value for the sample processed at 55 °C (58.95 g/100 g and 378.91 kcal/100 g, respectively) and low lipid and protein content (1.03 g/100 g and 1.39 g/100 g, respectively). Regardless of the sample analyzed, maltose was the majority sugar (37.33 - 281.01 g/100 g). The main fatty acids detected were oleic acid (61.33 - 62.08 %) followed by palmitic acid (33.91 - 34.40 %). The analysis of the mineral profile demonstrated that the samples did not differ significantly from each other, showing that the drying process did not interfere with the results obtained (p ≤ 0.05). The analysis of individual phenolics allowed the identification of six phenolic compounds in buriti shells. However, it is possible to observe that the drying method had a positive and significant influence on the phenolic profile (p ≤ 0.05), with chlorogenic acid (2.63 - 8.27 mg/100 g) and trigonelline (1.06 - 41.52 mg/100 g), the majority compounds. On the other hand, it is important to highlight that buriti shells have a high content of carotenoids, mainly ß-carotene (27.18 - 62.94 µg/100 g) and α-carotene (18.23 - 60.28 µg/100 g), also being positively influenced by the drying process at 55 °C (p ≤ 0.05). The dried shells showed a high content of phytochemical compounds and high antioxidant activity based on the different methods tested. The results show that buriti shell flour can be fully utilized and has nutritional and chemical aspects that can be applied to develop new sustainable, nutritious, and functional food formulations.

Stability of Bioactive Compounds and Antioxidant Activity in Rosehip Juice (Rosa spp.).

Rosehip fruits, characterized by their high concentrations of bioactive compounds and antioxidant activity (AA), have been traditionally used to make jams, infusions, and juices. Thus, the objective of this research was to evaluate the stability of rosehip juice by determining the concentrations of bioactive compounds and total phenols and the AA using chromatographic and spectroscopic methods. The stability of the juice was evaluated with three treatments and different storage conditions, namely, unpasteurized-refrigerated, pasteurized-room temperature, and pasteurized-refrigerated, and measurements were taken for eight months. Individual and total phenolic compounds, evaluated by chromatographic methods, reported differences until the end of this study. The total phenolic compounds by Folin-Ciocalteu method presented an average decrease of 57% in the three treatments in relation to the initial conditions. On the other hand, the ascorbic acid content decreased considerably, disappearing at week six. Furthermore, for the unpasteurized-refrigerated and pasteurized-refrigerated samples, a correlation was found between flavonols, total phenols, ascorbic acid, and antioxidant activity determined by the TEAC method. For the pasteurized-room temperature samples, correlations were found between the levels of several flavonols, hydroxycinnamic acid, total phenols, and ascorbic acid and the antioxidant activity determined by the CUPRAC method. The stability of the compounds was mainly correlated with the storage conditions of the juice and not with pasteurization. The highest stability was observed for the unpasteurized-refrigerated and pasteurized-refrigerated samples. Although the concentrations of the compounds evaluated decreased during this study, significant levels of AA persisted, providing beneficial characteristics for consumer health.

Validation of an Analytical Method of 3',4',5-Trihydroxy-3-Methoxy-6,7-Methylenedioxyflavone 4'-Glucuronide for Standardization of Spinacia oleracea.

Spinach (Spinacia oleracea) is one of the most famous vegetables worldwide, rich in essential metabolites for various health benefits. It is a valuable plant source that has the potential to be a nutraceutical. This study aimed to evaluate the single characteristic marker compound to establish the validation of HPLC-DAD methods applied to the development of a nutraceutical using spinach samples. Six metabolites (1-6) were identified from the spinach samples such as freeze-dried spinach (FDS) and spinach extract concentrate (SEC) by LC-Q-TOF/MS analysis. Among the six metabolites, 3',4',5-trihydroxy-3-methoxy-6,7-methylenedioxyflavone 4'-glucuronide (TMG) was selected as a marker compound due to its highest abundance and high selectivity. The specificity, accuracy, linearity, precision, repeatability, limit of detection (LOD), and limit of quantification (LOQ) of TMG in the spinach samples (FDS and SEC) were validated according to AOAC international guideline. The specificity was confirmed by monitoring the well separation of the marker compound from other compounds of spinach samples in the base peak intensity (BPI) and ultraviolet (UV) chromatogram. The calibration curve of TMG (15.625~500 µg/mL) had reasonable linearity (R2 = 0.999) considered with LOD and LOQ values, respectively. Recovery rate of TMG was 93-101% for FDS and 90-95% for SEC. The precision was less than 3 and 6% in the intraday and interday. As a result, the HPLC-DAD validation method of TMG in the spinach samples (FDS and SEC) was first established with AOAC and KFDA regulations for approving functional ingredients in functional foods.

A comprehensive approach to the monitoring of steroidal glycoalkaloids in foods of plant origin.

Steroidal glycoalkaloids (GAs) are toxins produced by solanaceous plants. As there are no fully standardized methods for their extraction and determination in food, the research aimed to: (1) develop and critically compare methods based on gas (GC) and liquid (LC) chromatography, including their coupling with mass spectrometry, and (2) to develop and optimize a universal GA extraction method. Hyphenated techniques proved to be the most useful in GA analysis: LC-MS was the most sensitive one, while GC-MS offered the highest chromatographic resolution. It was proven that quantitative results obtained using different analytical techniques cannot be directly compared. New extraction method that is more efficient than the AOAC method (997.13) was then designed and optimized. It was characterized by higher absolute recovery (99% and 34%, respectively) and allowed to extract much more GAs from the same material (e.g. 21.2 ± 1.4 and 11.82 ± 0.97 mg g-1 of potato tubers, respectively).