Chromatography-diode Array Detector-tandem Mass Spectrometry Research Articles

Cinnamomum sp. and Pelargonium odoratissimum as the Main Contributors to the Antibacterial Activity of the Medicinal Drink Horchata: A Study Based on the Antibacterial and Chemical Analysis of 21 Plants.

Horchata, a herbal infusion drink from Ecuador containing a mixture of medicinal plants, has been reported to exhibit anti-inflammatory, analgesic, diuretic, and antioxidant activity. The antibacterial activity of each of the plants contained in the horchata mixture has not been fully evaluated. Thus, in this study, we analysed the antibacterial activity of 21 plants used in horchata, collected from the Ecuadorian Andes region, against bacterial strains of clinical importance. The methanolic extract of Cinnamomum sp. showed minimal inhibitory concentration (MIC) values of 250 µg/mL against Staphylococcus aureus ATCC25923 and Methicillin-resistant S. aureus (MRSA), while Pelargonium odoratissimum exhibited a MIC value of 500 µg/mL towards S. aureus ATCC25923. The high-performance liquid chromatography-diode array detector-tandem mass spectrometry (HPLC-DAD-MS/MS) analyses identified in Cinnamomum sp. epicatechin tannins, cinnamaldehyde, and prehelminthosporol molecules, whereas in P. odoratissimum, gallocatechin and epigallocatechin tannins, some flavonoids, and gallic acid and derivatives were identified. Finally, Cinnamomum sp. and P. odoratissimum showed partial inhibition of biofilm formation of S. aureus ATCC25923 and MRSA. Overall, our findings revealed which of the plants used in horchata are responsible for the antibacterial activity attributed to this herbal drink and exhibit the potential for Cinnamomum sp. and P. odoratissimum secondary metabolites to be explored as scaffolds in drug development.

Steryl ferulates composition in twenty-two millet samples: Do “microwave popping” and fermentation affect their content?

Climate change has led to rediscovery of minor drought tolerant grains such as millet. Among its bioactive molecules, steryl ferulates have been poorly explored. Steryl ferulates composition of was investigated by high performance liquid chromatography-diode array detector-tandem mass spectrometry and high resolution tandem mass spectrometry in twenty-two millet samples and also in some fermented and microwave heated samples. Six compounds were found in Panicum, Pennisetum, Eleusine and Setaria genera, with a prevalence of campestanyl and sitostanyl ferulate. The lowest steryl ferulates content was found in Panicum, with values ranging from 2.98 ± 0.04 µg/g to 8.72 ± 0.41 µg/g. Foxtail millet and finger millet showed the highest amount with 46.07 ± 5.20 µg/g and 85.29 ± 4.30 µg/g, respectively. As for pearl millet, microwave heating and fermentation increased steryl ferulates by two (33.77 ± 0.88 µg/g) and five (75.83 ± 1.25 µg/g) times, with respect to the untreated sample. Microwave heating and fermentation could be used to increase steryl ferulates in millet.

Chemical Characterisation, Antioxidant and Antibacterial Activities of Pinus pinaster Ait. and Pinus pinea L. Bark Polar Extracts: Prospecting Forestry By-Products as Renewable Sources of Bioactive Compounds

Agroforestry by-products have gained rising attention in recent years as they represent inexpensive and abundant raw materials that are a source of added-value chemicals, e.g., for food and pharmaceutical applications, as well as for bioenergy generation. Pinus pinaster Ait. bark extracts are consumed worldwide for their cardiovascular benefits, whilst the health potential of Pinus pinea L. bark has not yet been deeply exploited. Therefore, this study highlights the chemical characterisation of Portuguese P. pinaster Ait. and P. pinea L. bark polar extracts, via ultra-high performance liquid chromatography-diode array detection-tandem mass spectrometry (UHPLC-DAD-MSn) analysis, and their antioxidant and antibacterial activities. Quinic acid, an A-type procyanidin dimer isomer, protocatechuic acid, and quercetin were identified for the first time as P. pinea L. bark components. Moreover, this bark demonstrated a higher total content of identified polar compounds than P. pinaster Ait. bark, with quinic acid being the most abundant compound identified. Regarding antioxidant activity, the pine bark polar extracts exhibited strong reducing power and 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2-azinobis-(3-ethyl-benzothiazoline-6-sulfonic acid (ABTS) radical scavenging effects compared to natural antioxidants. Moreover, the bactericidal actions of pine bark extracts were shown against Staphylococcus aureus and Escherichia coli at a 3.13–25 mg mL−1 range. Globally, these promising insights can boost the sustainable exploitation of P. pinea L. bark, as already occurs with P. pinaster Ait. bark, for the food and biomedical fields.

Antimicrobial efficacy and biocompatibility of extracts from Cryptocarya species.

This study evaluated the efficacy of Cryptocarya spp extracts on biofilm of Candida albicans and its biocompatibility. Mature biofilm of C. albicans was formed on denture base acrylic resin samples and the fungicidal effect of the extracts was evaluated by Alamar Blue® assay, counting colony-forming units (CFU/mL) and confocal laser scanning microscopy (CLSM). Cytotoxicity of extracts from Cryptocarya species was evaluated by AlamarBlue® assay, using normal oral keratinocytes (NOK) cells. In additional, Analysis of plant extracts by ultra-high-performance liquid chromatography-diode array detector-tandem mass spectrometry (UPLC-DAD-MS) was performed. The results showed significant reduction in the cellular metabolism and in the number of CFU/mL of C. albicans (p<0.05). The concentration of 0.045 g/mL completely inhibited the number of CFU/mL. Regarding cytotoxicity, all extracts decreased cell viability compared to the control group. CLSM analysis showed predominance of live cells, but with a great difference between the groups. Antimicrobial activity of extracts from Cryptocarya on C. albicans biofilm was confirmed. However, all extracts showed toxicity on NOK cells.

Anti-Inflammatory Effects of Jakyakgamcho-Tang in IL-4- and TNF-α-Stimulated Lung Epithelial Cells and Lipopolysaccharide-Stimulated Macrophages

Asthma is a chronic respiratory disease mediated by airway inflammation. Jakyakgamcho-tang (JGT), a traditional medicine, is widely subscribed to common diseases such as muscle pain and cramps in East Asian countries. Although the efficacy of JGT on peripheral neuropathy, gouty arthritis, and colitis has been reported, the effect of JGT on airway inflammation related to asthma is not clearly investigated. In this study, we aimed to evaluate the effects of JGT water extract (JGTW) on factors related to airway inflammation using the human bronchial epithelial BEAS-2B and the mouse monocyte-macrophage RAW264.7 cell lines. Furthermore, the constituents in JGTW were quantitatively and qualitatively studied for future reference of JGTW standardization. JGTW reduced the generation of several airway inflammation mediators such as eotaxins, regulated on activation normal T-cell expressed and secreted (RANTES), and matrix metalloproteinase-9, and expressions of adhesion molecules (ICAM-1 and VCAM-1), which attracts leukocytes to the site of inflammation in interleukin-4 + tumor necrosis factor-α (IT)-stimulated BEAS-2B cells. In lipopolysaccharide-stimulated RAW264.7 cells, JGTW effectively suppressed inducible nitric oxide synthase (iNOS) induction by inhibiting the MAPK and NF-κB signaling. In addition, JGTW treatment showed decreased inflammatory cells and Th2 cytokines in bronchoalveolar lavage fluid and decreased IgE levels in plasma in the OVA-induced asthmatic mice model. In the ultra-performance liquid chromatography-diode array detector-tandem mass spectrometry analysis, 24 phytochemicals were identified in JGTW, and paeoniflorin (63.971 mg/g) and glycyrrhizin (11.853 mg/g) were found to be the most abundant. These findings suggest that JGTW has anti-inflammatory effects on airway inflammation by regulating inflammatory response-related factors, possibly through MAPK and NF-κB in pulmonary epithelial cells and macrophages.

Investigation of Praziquantel/Cyclodextrin Inclusion Complexation by NMR and LC-HRMS/MS: Mechanism, Solubility, Chemical Stability, and Degradation Products

Praziquantel (PZQ) is a biopharmaceutical classification system (BCS) class II anthelmintic drug characterized by poor solubility and a bitter taste, both of which can be addressed by inclusion complexation with cyclodextrins (CD). In this work, a comprehensive investigation of praziquantel/cyclodextrin (PZQ/CD) complexes was conducted by means of UV–vis spectroscopy, spectrofluorimetry, NMR spectroscopy, liquid chromatography-high-resolution mass spectrometry (LC-HRMS/MS), and molecular modeling. Phase solubility studies revealed that among four CDs tested, the randomly methylated β-CD (RMβCD) and the sulfobutylether sodium salt β-CD (SBEβCD) resulted in the highest increase in PZQ solubility (approximately 16-fold). The formation of 1:1 inclusion complexes was confirmed by HRMS, NMR, and molecular modeling. Both cyclohexane and the central pyrazino ring, as well as an aromatic part of PZQ are included in the CD central cavity through several different binding modes, which exist simultaneously. Furthermore, the influence of CDs on PZQ stability was investigated in solution (HCl, NaOH, H2O2) and in the solid state (accelerated degradation, photostability) by ultra-high-performance liquid chromatography–diode array detection–tandem mass spectrometry (UPLC-DAD/MS). CD complexation promoted new degradation pathways of the drug. In addition to three already known PZQ degradants, seven new degradation products were identified (m/z 148, 215, 217, 301, 327, 343, and 378) and their structures were proposed based on HRMS/MS data. Solid complexes were prepared by mechanochemical activation, a solvent-free and ecologically acceptable method.

Clitorienolactones and Isoflavonoids of Clitorea ternatea Roots Alleviate Stress-Like Symptoms in a Reserpine-Induced Zebrafish Model.

Clitorea ternatea has been used in Ayurvedic medicine as a brain stimulant to treat mental illnesses and mental functional disorders. In this study, the metabolite profiles of crude C. ternatea root extract (CTRE), ethyl acetate (EA), and 50% aqueous methanol (50% MeOH) fractions were investigated using ultrahigh-performance liquid chromatography–diode array detector–tandem mass spectrometry (UHPLC–DAD–MS/MS), while their effect on the stress-like behavior of zebrafish, pharmacologically induced with reserpine, was investigated. A total of 32 compounds were putatively identified, among which, a series of norneolignans, clitorienolactones, and various flavonoids (flavone, flavonol, isoflavone, and isoflavanone) was found to comprise the major constituents, particularly in the EA and 50% MeOH fractions. The clitorienolactones, presently unique to the species, were present in both the free and glycosylated forms in the roots. Both the EA and 50% MeOH fractions displayed moderate effects on the stress-induced zebrafish model, significantly decreasing freezing duration and elevating the total distance travelled and average velocity, 72 h post-treatment. The results of the present study provide further evidence that the basis for the use of C. ternatea roots in traditional medicine to alleviate brain-related conditions, such as stress and depression, is attributable to the presence of clitorienolactones and the isoflavonoidal constituents.

Inhibitory Effects of Gyeji-Tang on MMP-9 Activity and the Expression of Adhesion Molecules in IL-4- and TNF-α-Stimulated BEAS-2B Cells.

Gyeji-tang (GJT), a traditional herbal formula composed of five herbal medicines, is commonly used to treat the common cold, exogenous febrile disease, fever and headaches in Korea, China and Japan. Although various pharmacological activities of GJT have been reported in several studies, the effect of GJT water extract (GJTWE) on airway inflammation has not yet been investigated. This study aimed to evaluate the effects of GJTWE on airway inflammation-related factors using human bronchial epithelial BEAS-2B cells, and to identify the phytochemicals in GJTWE by ultra-performance liquid chromatography-diode array detector-tandem mass spectrometry (UPLC-DAD-MS/MS) analysis. GJTWE significantly decreased the production of chemokines, including eotaxin-3, eotaxin-1, regulated on activation normal T-cell expressed and secreted (RANTES), and matrix metalloproteinase-9, and the expression of the adhesion molecules, intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, in interleukin-4 + tumor necrosis factor-α (IT)-stimulated BEAS-2B cells. In the UPLC-DAD-MS/MS analysis, 21 phytochemicals, including six flavonoids, two chalcones, five terpenoids, six phenolics, one phenylpropanoid and one coumarin, were identified in GJTWE. The findings suggested that GJTWE might exhibit anti-inflammatory effects on airway inflammation by regulating the expression of inflammatory response-related factors in IT-stimulated BEAS-2B cells; further studies are required to determine the bioactive compounds involved in the inhibition of airway inflammation.

Modification of Natural Proanthocyanidin Oligomers and Polymers Via Chemical Oxidation under Alkaline Conditions.

We tested the susceptibility of 102 proanthocyanidin (PA)-rich plant extracts to oxidation under alkaline conditions and the possibility to produce chemically modified PAs via oxidation. Both the nonoxidized and the oxidized extracts were analyzed using group-specific ultrahigh-performance liquid chromatography–diode array detection–tandem mass spectrometry (UHPLC–DAD–MS/MS) methods capable of detecting procyanidin (PC) and prodelphinidin (PD) moieties along the two-dimensional (2D) chromatographic fingerprints of plant PAs. The results indicated different reactivities for PCs and PDs. When detected by UHPLC–DAD only, most of the PC-rich samples exhibited only a subtle change in their PA content, but the UHPLC–MS/MS quantitation showed that the decrease in the PC content varied by 0–100%. The main reaction route was concluded to be intramolecular. The PD-rich and galloylated PAs showed a different pattern with high reductions in the original PA content by both ultraviolet (UV) and MS/MS quantitation, accompanied by the shifted retention times of the chromatographic PA humps. In these samples, both intra- and intermolecular reactions were indicated.

Formulation of Novel Liqueurs from Juice Industry Waste: Consumer Acceptance, Phenolic Profile and Preliminary Monitoring of Antioxidant Activity and Colour Changes During Storage.

SUMMARYResearch backgroundApple juice is one of the most popular and liked beverages worldwide. Due to the increased health consciousness among consumers, beetroot and chokeberry juices have also rising consumption trends. Despite representing a considerable percentage of the processed fruit and rich source of bioactive compounds, fruit pomace, remaining after juice production, has still been underutilised. Here, the possibility of using apple, beetroot and chokeberry pomace in liqueur formulations is investigated.Experimental approachApple and chokeberry liqueurs were produced from apple and chokeberry pomace extracts, respectively. Apple/chokeberry and apple/beetroot liqueurs were obtained by combining apple pomace with chokeberry and beetroot pomace extracts in ratios 50:50 and 70:30, respectively. The sensory quality and acceptability of freshly prepared liqueurs were evaluated by experts and consumers. Sugars and phenolics were identified and quantified by high-performance anion-exchange chromatography with pulsed-amperometric detection (HPAEC-PAD) and high-performance liquid chromatography–diode array detection–tandem mass spectrometry (HPLC–DAD–MS/MS), respectively. Storability was preliminarily evaluated based on monitoring of total phenolic concentration, antioxidant activity and colour each month during 6 months of storage at 4 and 22 °C.Results and conclusionsThe expert and the consumer testing indicated that apple and chokeberry pomace could be used as raw materials without any flavour corrections while apple/beetroot pomace liqueur would require modification. High total phenolic content and antioxidant activity were found in all freshly prepared liqueurs, with chokeberry liqueur being by far superior. Among identified phenolics, ellagic acid and phlorizin were quantified as the most prominent, except in chokeberry liqueur, where phlorizin was not quantified. Despite the decrease in total phenolic concentration and antioxidant activity after 6 months, liqueurs still represented a rich source of phytochemicals. The highest phenolic compound retention and antioxidant activity maintenance were observed in chokeberry liqueur. Also, the appealing colour was retained despite the changes detected in chromatic characteristics.Novelty and scientific contributionThe possibility of apple, beetroot and chokeberry pomace restoration into the food chain by the production of liqueurs has been demonstrated for the first time. Functional and sensorial properties of newly developed liqueurs indicated that the selected pomace represents the promising raw material for liqueur production. The applied approach represents a contribution to the circular economy in juice production.

Miracle Fruit Leaf Extract: Antioxidant Activity Evaluation, Constituent Identification, and Medical Applications

A systematic study was carried out on the antioxidants in miracle fruit leaf extract. The total phenols, total flavonoids, saponins and the antioxidant activities of the extract were evaluated using the 2,2′-diphenyl-2-picrylhydrazyl (DPPH), 2,2′-azino-bis-3-ethyl-benzotiazolin-6-sulfonic acid (ABTS), metal chelating capacity (MCC) and ferric reducing antioxidant power (FRAP) assays. The results showed that the leaves contained high phenol concentrations of 90.0 ± 4.78 mg gallic acid equivalent/g dry weight, total flavonoids at 33.0 ± 1.92 mg rutin equivalent/g dry weight, and saponins at 27.8 ± 1.64 mg oleanolic acid equivalent/g dry weight. The extract possessed excellent free radical scavenging ability and high antioxidant capacities. Furthermore, eighteen polyphenol and flavonoid constituents in the extract were identified by using ultra-high performance liquid chromatography-diode array detector-tandem mass spectrometry (UPLC-DAD-MSn) method and standard chemicals. The constituents were gallic acid, 5-O-caffeoylquinic acid, 3-O-p-coumaroylquinic acid, 5-O-p-coumaroylquinic acid, 3-O-feruloylquinic acid, 4-O-caffeoylquinic acid, 4-O-p-coumaroylquinic acid dimer, 4-O-p-coumaroylquinic acid, 4-O-feruloylquinic acid, 4-O-feruloylquinic acid, rutin, myricetin-3-O-rhamnoside, quercetin-3-D-galactoside, quercetin-3-glucoside, kaempferol-3-O-glucoside, quercetin-3-rhamnoside, myricetin-O-galloylrhamnoside and quercetin-3-O-(2′′-galloyl)-rhamnoside.Eight constituents were quantified with values from 0.12 to 4.13 mg·g‒1. The extract of miracle fruit leaves was also used in an anti-Alzheimer's disease model for potential medical applications. The assay indicated that the extract provided apparent inhibition for the paralysis of Caenorhabditis elegans at a concentration of 0.02 mg·mL‒1. This work demonstrated that miracle fruit leaves are rich in bioactive antioxidants with potential for drug and functional product development.

Environmentally Friendly Hydrothermal Processing of Melon by-Products for the Recovery of Bioactive Pectic-Oligosaccharides.

Melon by-products, that currently lack high value-added applications, could be a sustainable source of bioactive compounds such as polysaccharides and antioxidants. In this work, melon peels were extracted with water to remove free sugars, and the water-insoluble solids (WISs) were subjected to hydrothermal processing. The effect of temperature on the composition of the obtained liquors and their total phenolic content was evaluated. The selected liquors were also characterized by matrix assisted laser desorption/ionization-time of flight mass spectroscopy (MALDI-TOF MS), fourier transform infrared spectroscopy (FTIR) and high performance anion exchange chromatography with pulsed amperometric detection (HPAEC–PAD), and its phenolic compounds were identified and quantified by high-performance liquid chromatography-diode array detector-tandem mass spectrometry (HPLC–DAD–MS/MS). In addition, the spent solids from the hydrothermal treatment were characterized and their potential use was assessed. At the optimal conditions of 140 °C (severity 2.03), the total oligosaccharide yield accounted for 15.24 g/100 g WIS, of which 10.07 g/100 g WIS were oligogalacturonides. The structural characterization confirmed the presence of partially methyl esterified oligogalacturonides with a wide range of polymerization degrees. After precipitation, 16.59 g/100 g WIS of pectin were recovered, with a galacturonic acid content of 55.41% and high linearity.

Therapeutic Effects of Morinda citrifolia Linn. (Noni) Aqueous Fruit Extract on the Glucose and Lipid Metabolism in High-Fat/High-Fructose-Fed Swiss Mice.

The aim of this study was to evaluate the therapeutic effects of two different doses (250 and 500 mg/kg) of Morinda citrifolia fruit aqueous extract (AE) in high-fat/high-fructose-fed Swiss mice. The food intake, body weight, serum biochemical, oral glucose tolerance test (OGTT), and enzyme-linked immunosorbent assay (ELISA), as well as histological analyses of the liver, pancreatic, and epididymal adipose tissue, were used to determine the biochemical and histological parameters. The chemical profile of the extract was determined by ultra-fast liquid chromatography–diode array detector–tandem mass spectrometry (UFLC–DAD–MS), and quantitative real-time PCR (qRT-PCR) was used to evaluate the gene expressions involved in the lipid and glucose metabolism, such as peroxisome proliferative-activated receptors-γ (PPAR-γ), -α (PPAR-α), fatty acid synthase (FAS), glucose-6-phosphatase (G6P), sterol regulatory binding protein-1c (SREBP-1c), carbohydrate-responsive element-binding protein (ChREBP), and fetuin-A. Seventeen compounds were tentatively identified, including iridoids, noniosides, and the flavonoid rutin. The higher dose of AE (AE 500 mg/kg) was demonstrated to improve the glucose tolerance; however, both doses did not have effects on the other metabolic and histological parameters. AE at 500 mg/kg downregulated the PPAR-γ, SREBP-1c, and fetuin-A mRNA in the liver and upregulated the PPAR-α mRNA in white adipose tissue, suggesting that the hypoglycemic effects could be associated with the expression of genes involved in de novo lipogenesis.

Optimisation of Sequential Microwave-Assisted Extraction of Essential Oil and Pigment from Lemon Peels Waste.

In this work, a cascade approach to obtain different valuable fractions from lemon peels waste was optimised using microwave-assisted processes. Microwave-assisted hydrodistillation (MAHD) with a Clevenger apparatus was firstly used to obtain the lemon essential oil (LEO). The remaining residue was then submitted to microwave-assisted extraction (MAE) to extract the lemon pigment (LP). A Box–Behnken design was used to evaluate the influence of ethanol concentration, temperature and time in LP extraction in terms of extraction yield and colour intensity. Optimal extraction conditions for LP were 80% (v/v) ethanol, 80 °C and 50 min, with a liquid-to-solid ratio of 1:10. The obtained yields for LEO and LP were around 2 wt.% and 6 wt.%, respectively. The composition of LEO was analysed by gas chromatography with flame ionisation detection (GC-FID), and limonene (65.082 wt.%), β-pinene (14.517 wt.%) and γ-terpinene (9.743 wt.%) were mainly identified. LP was purified by using different Amberlite adsorption resins (XAD4, XAD7HP and XAD16N), showing XAD16N the best adsorption capacity. Enrichment factors of 4.3, 4.5 and 5.0 were found for eriocitrin, diosmin and hesperidin, respectively, which were detected as the main components in LP by ultra-high-performance liquid chromatography–diode array detector–tandem mass spectrometry (UPLC-DAD-MS) analysis, with final concentrations of 4.728 wt.%, 7.368 wt.% and 2.658 wt.%, respectively. Successful antimicrobial capacity against Escherichia coli and Staphylococcus aureus was obtained for LEO. The results from this work showed the potential of applying a cascading approach based on microwave-assisted processes to valorise lemon wastes, obtaining natural pigments and antimicrobials to be applied in food, cosmetic and polymer industries.

A HPLC-DAD-MS/MS Method for Simultaneous Determination of Six Active Ingredients of Salviae Miltiorrhizae and Ligustrazine Hydrochloride Injection in Rat Plasma and its Application to Pharmacokinetic Studies.

This is a pharmacokinetic study of Salviae miltiorrhizae and ligustrazine hydrochloride injection. The study aimed to evaluate the mechanism of action, safety and rational clinical use of Salviae miltiorrhizae and ligustrazine hydrochloride injection. Salviae miltiorrhizae and ligustrazine hydrochloride injection is a compound preparation consisted of Salvia miltiorrhiza extract and ligustrazine hydrochloride for the treatment of cardiovascular and cerebrovascular diseases in China. The study aimed to develop a rapid and sensitive high-performance liquid chromatography-diode array detector-tandem mass spectrometry (HPLC-DAD-MS/MS) method for simultaneous determination of six major active ingredients of Salviae miltiorrhizae and ligustrazine hydrochloride injection, namely danshensu, protocatechuic aldehyde, rosmarinic acid, lithospermic acid, salvianolic acid A, and ligustrazine hydrochloride, in rat plasma. Plasma samples were precipitated with methanol, which was spiked with ascorbic acid and the supernatant was separated on a Waters Cortecs C18 column, by using a gradient mobile phase system of acetonitrile-water containing 0.05% formic acid (v/v). For internal standards, puerarin was selected for the five salvianolic acids, while isofraxidin was used for ligustrazine hydrochloride. Besides, electrospray ionization in negative mode and multiplereaction monitoring were used to identify and quantify the five salvianolic acids, whereas ligustrazine hydrochloride was quantified at 310 nm using the diode array detector. Noticeably, all calibration curves showed good linearity (R2>0.99) over the concentration range, with a lower limit of quantification between 0.00411 and 0.0369 μg/mL for salvianolic acids, and 1.74 μg/mL for ligustrazine hydrochloride. Next, the precision of the developed method was evaluated by intra- and inter-day assays, and the percentage of relative standard deviation was within 10%. Although the extraction efficiency of some salvianolic acids was not very satisfactory, the sensitivity of the analytical method met the analysis requirements of rat plasma samples. Moreover, the validated method was successfully applied to a pharmacokinetic study of Salviae miltiorrhizae and ligustrazine hydrochloride injection in the rat model. Linear pharmacokinetic characteristics were observed for the six active ingredients after intravenous infusion administration in rats within the dose range examined here. In summary, our study proposed a HPLC-DADMS/ MS method with the simultaneous determination of multiple ingredients, and demonstrated its applicability in pharmacokinetic studies.

Spectrophotometric determination of the antioxidant properties and characterization of the phenolic content by high-performance liquid chromatography–diode array detection–tandem mass spectrometry (HPLC–DAD–MS/MS) of Berberis hispanica Boiss. & Reut. leaves

Here is reported the phenolic profile obtained by high-performance liquid chromatography with diode array and mass spectrometry detection and the antioxidant activity using the ABTS and DPPH assays of leaves of Berberis hispanica Boiss. & Reut. The most abundant compounds were caffeoylquinic acids, in particular chlorogenic acid with 19.3 mg g−1 dried extract, caffeoylglucaric acids and quercetin glycosides. Also, the compound dihydro-roseoside is reported for the first time in a Berberis species. The antioxidant potential was in agreement with the phenolic content: the highest concentration of phenolics corresponded to the highest antioxidant activity in both of the assays equal to 648 and 547 µmol Trolox equivalents g−1 dried extract methanolic fraction for the ABTS and DPPH assays, respectively. These results contribute to the limited bibliographic information concerning this plant species and represent a starting point to evaluate its potential as a novel source of bioactive compounds.

Myrtle Seeds (Myrtus communis L.) as a Rich Source of the Bioactive Ellagitannins Oenothein B and Eugeniflorin D2.

The increasing popularity of “Mirto” liqueur, produced from Myrtus communis berries, has led to the planting of domesticated cultivars, expanding myrtle berry production. To promote the use of cultivated berries, the content in the nutraceutical compounds ellagitannins has been investigated both in spontaneous and cultivated fruits. Oenothein B and eugeniflorin D2, characterized by 1H and 13C NMR, were isolated and quantified using ultrahigh-performance liquid chromatography–diode array detector–tandem mass spectrometry (UPLC–DAD–MS/MS). The antifungal and anti-inflammatory activities of oenothein B were assayed in vitro. Large amounts of oenothein B and eugeniflorin D2 were detected in seeds (12 ± 2.4 and 5.8 ± 1.2 mg/g). The oenothein B concentration in liqueurs was 194 ± 22 mg/L. This macrocyclic ellagitannin dimer showed anti-Candida (minimal inhibitory concentration <8–64 μg/mL) and anti-inflammatory properties. Cultivated myrtle berries are a source of nutraceutical compounds. The high concentration of oenothein B in liqueur suggests a possible contribution to the organoleptic and biological properties of the beverage.

Chemical Fingerprint of 'Oblačinska' Sour Cherry (Prunuscerasus L.) Pollen.

The aim of this research was to analyze sugars and phenolics of pollen obtained from 15 different ‘Oblačinska’ sour cherry clones and to assess the chemical fingerprint of this cultivar. Carbohydrate analysis was done using high-performance anion-exchange chromatography (HPAEC) with pulsed amperometric detection (PAD), while polyphenols were analyzed by ultra-high-performance liquid chromatography–diode array detector–tandem mass spectrometry (UHPLC-DAD MS/MS) system. Glucose was the most abundant sugar, followed by fructose and sucrose. Some samples had high level of stress sugars, especially trehalose. Rutin was predominantly polyphenol in a quantity up to 181.12 mg/kg (clone III/9), with chlorogenic acid (up to 59.93 mg/kg in clone III/9) and p-coumaric acid (up to 53.99 mg/kg in clone VIII/1) coming after. According to the principal component analysis (PCA), fructose, maltose, maltotriose, sorbitol, and trehalose were the most important sugars in separating pollen samples. PCA showed splitting off clones VIII/1, IV/8, III/9, and V/P according to the quantity of phenolics and dissimilar profiles. Large differences in chemical composition of studied ‘Oblačinska sour cherry’ clone pollen were shown, proving that it is not a cultivar, but population. Finally, due to the highest level of phenolics, clones IV/8, XV/3, and VIII/1 could be singled out as a promising one for producing functional food and/or in medicinal treatments.

Optimization of Ultrasound-Assisted Extraction of Antioxidant Polyphenols from the Seed Coats of Red Sword Bean (Canavalia gladiate (Jacq.) DC.).

The seed coat of red sword bean (Canavalia gladiata (Jacq.) DC.) is rich in antioxidant polyphenols. It is often discarded as a byproduct with the consumption of red sword bean, since it is very thick and not consumed by people. The aim of this study was to develop an ultrasound-assisted extraction method to extract natural antioxidants from the seed coats. The extraction process was optimized by using response surface methodology. After the single-factor experiments, three key factors, including ethanol concentration, liquid/solid ratio, and extraction time, were selected and their interactions were studied using a central composite design. The optimal extraction condition was 60.2% hydroethanol, a liquid/solid ratio of 29.3 mL/g, an extraction time of 18.4 min, an extraction temperature of 50 °C, and ultrasound power of 400 W. Under the optimal conditions, antioxidant activity of the extract was 755.98 ± 10.23 μmol Trolox/g dry weight (DW), much higher than that from maceration (558.77 ± 14.42 μmol Trolox/g DW) or Soxhlet extraction (479.81 ± 12.75 μmol Trolox/g DW). In addition, the main antioxidant compounds in the extract were identified and quantified by high-performance liquid chromatography–diode array detection–tandem mass spectrometry (HPLC–DAD–MS/MS). The concentrations of digalloyl hexoside, methyl gallate, gallic acid, trigalloyl hexoside, and digallic acid were 15.30 ± 0.98, 8.85 ± 0.51, 8.76 ± 0.36, 4.27 ± 0.21, and 2.89 ± 0.13 mg/g DW. This study provides an efficient and green extraction method for the extraction of natural antioxidants from the bean coat of red sword bean. The extract of antioxidants might be added into functional foods or nutraceuticals with potential beneficial functions.

Preventive Effects of an UPLC-DAD-MS/MS Fingerprinted Hydroalcoholic Extract of Citrus aurantium in a Mouse Model of Ulcerative Colitis.

Although traditionally used to improve indigestion, diarrhea, dysentery, and constipation, the therapeutic effects of Citrus aurantium on intestinal inflammation remain unclear. The aim of this study was to evaluate the beneficial effects and to identify the active components of a hydroalcoholic extract of C. aurantium (HECA) on ulcerative colitis. HECA was prepared with 70% ethanol solution in water and extracted at 37 °C for 12 h in triplicate, filtered through a sieve, and lyophilized. Phytochemical identification of HECA was performed by ultra-performance liquid chromatography-diode array detector-tandem mass spectrometry (UPLC-DAD-MS/MS). Animals were randomly assigned to one of four groups based on the treatment conditions. Ulcerative colitis was induced by administration of 2.5% dextran sodium sulfate (DSS) in drinking water for 5 d. Body weight, clinical signs, colon length, pro-inflammatory cytokine expression levels, and histopathological findings were evaluated. In UPLC-DAD-MS/MS analysis, the identified phytochemical components of HECA included four alkaloids, seven coumarins, 18 flavonoids, two lignans, two phenolics, and 10 terpenoids. HECA markedly protected against body weight loss and colon shortening. In pathological examination, HECA alleviated DSS-related mucosal inflammatory lesions in the colon. Moreover, HECA markedly reduced the expression levels of interleukin-6, interferon-γ, tumor necrosis factor-α, and monocyte chemotactic protein-1 in colonic inflammation. Taken together, HECA has potential to relieve mucosal inflammation in the colon, suggesting that the putative active ingredients are responsible for the anti-ulcerative effects.