Quantification Of Compounds Research Articles

Rapid and simultaneous quantification of phenolic compounds in peanut (Arachis hypogaea L.) seeds using NIR spectroscopy coupled with multivariate calibration

Phytochemically, peanuts provide significant nutritional value to humans, animals, and the food industry as a whole. This study was conducted to explore the potential for near-infrared (NIR) spectroscopy and effective variable selection algorithms to quantify phenolic compounds in peanut seeds. The phenolics were extracted and then identified and quantified using high-performance liquid chromatography (HPLC). The spectroscopic data were acquired from the peanut samples using a tabletop NIR spectrometer with a wavelength range of 10,000–4000 cm–1. The acquired spectra were preprocessed using a synergistic effect of first- and second-order derivative (FOD and SOD) preprocessing techniques, and multivariate algorithms were used, examined, and evaluated using correlation coefficients of the validation set (Rp), root mean square error of prediction (RMSEP), and residual predictive deviations (RPDs). The competitive adaptive reweighted sampling-partial least squares (CARS-PLS) model produced optimum performance for chlorogenic acid (Rp = 0.933, RPD = 2.77), kaempferol (Rp = 0.928, RPD = 2.68), p-coumaric acid (Rp = 0.900, RPD = 2.32), and quercetin (Rp = 0.932, RPD = 2.88), respectively. Therefore, this study proved that NIR spectroscopy in combination with CARS-PLS was capable of nondestructively predicting phenolic content in peanut seeds.

Quantification of pharmaceutical compounds in tissue and plasma samples using selective ion accumulation with multiple mass isolation windows.

Quantification of pharmaceutical compounds using matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is an alternative to traditional liquid chromatography (LC)-MS techniques. Benefits of MALDI-based approaches include rapid analysis times for liquid samples and imaging mass spectrometry capabilities for tissue samples. As in most quantification experiments, the use of internal standards can compensate for spot-to-spot and shot-to-shot variability associated with MALDI sampling. However, the lack of chromatographic separation in traditional MALDI analyses results in diminished peak capacity due to the chemical noise background, which can be detrimental to the dynamic range and limit of detection of these approaches. These issues can be mitigated by using a hybrid mass spectrometer equipped with a quadrupole mass filter (QMF) that can be used to fractionate ions based on their mass-to-charge ratios. When the masses of the analytes and internal standards are sufficiently disparate in mass, it can be beneficial to effect multiple narrow mass isolation windows using the QMF, as opposed to a single wide mass isolation window, to minimize chemical noise while allowing for internal standard normalization. Herein, we demonstrate a MALDI MS quantification workflow incorporating multiple sequential mass isolation windows enabled on a QMF, which divides the total number of MALDI laser shots into multiple segments (i.e., one segment for each mass isolation window). This approach is illustrated through the quantitative analysis of the pharmaceutical compound enalapril in human plasma samples as well as the simultaneous quantification of three pharmaceutical compounds (enalapril, ramipril, and verapamil). Results show a decrease in the limit of detection, relative standard deviations below 10%, and accuracy above 85% for drug quantification using multiple mass isolation windows. This approach has also been applied to the quantification of enalapril in brain tissue from a rat dosed in vitro. The average concentration of enalapril determined by imaging mass spectrometry is in agreement with the concentration determined by LC-MS, giving an accuracy of 104%.

Simultaneous identification and quantification of pentacyclic triterpenoids and phenolic compounds from the leaves of Boswellia serrata using LC-MS/MS tandem mass spectrometry.

Boswellia serrata (B. serrata) is an important medicinal plant widely used as dietary supplements to provide a support for osteoarthritic and inflammatory diseases. The occurrence of triterpenes in leaves of B. serrata is very little or none. Therefore, the qualitative and quantitative determination of phytoconstituents (triterpenes and phenolics) present in the leaves of B. serrata is very much needed. The aim of this study was to develop an easy, rapid, efficient and simultaneous liquid chromatography-mass spectrometry (LC-MS/MS) method for the identification and quantification of the compounds present in the leaves extract of B. serrata. The purification of ethyl acetate extracts of B. serrata was performed by solid phase extraction method, followed by HPLC-ESI-MS/MS analysis. Chromatographic parameters of the analytical method included negative electrospray ionization (ESI-) with a flow of 0.5mL/min in gradient mode consisting of acetonitrile (A) and water (B) containing 0.1% formic acid, at 20°C. Total 19 compounds (13 triterpenes and 6 phenolic compounds) were separated, and simultaneously quantified using a validated LC-MS/MS method with high accuracy and sensitivity. Good linearity was obtained with r2 > 0.973 in the calibration range. The overall recoveries were in a range between 95.78 and 100.2% with relative standard deviations (RSD) below 5% for the entire procedure of matrix spiking experiments. Overall, there was no ion suppression from the matrix. The quantification data showed that the total amount of triterpenes and phenolic compounds in the leaves of B. serrataethyl acetate extract samples ranged from 14.54 to 102.14mg/g and 2.14 to 93.12mg/g of dry extract, respectively. This work provides, for the first time, a chromatographic fingerprinting analysis on the leaves of B. serrata. A rapid, efficient, and simultaneous liquid chromatography-mass spectrometry (LC-MS/MS) method was developed and used for the both identification and quantification of triterpenes and phenolic compounds in the leaves extracts of B. serrata. The method established in this work can be used as quality-control method for other market formulations or dietary supplements containing leaf extract of B. serrata.

Response Surface Methodology as an Experimental Strategy for Ultrasound-Assisted Extraction of Phenolic Compounds from Artichoke Heads.

The accurate quantification of phenolic compounds (PCs) in foods has become mandatory for a reliable estimation of PCs dietary intake. However, the extraction step of these molecules from the food matrix is a challenging and complex task. To manage the current lack of an official or generally accepted procedure for the recovery of phenolics, the application of statistical and mathematical tools, such as the response surface methodology (RSM), that allow the optimization of extraction parameters and the acquisition of the best output, has become the analytical approach of choice. The aim of this study was to apply an RSM-optimized ultrasound-assisted procedure to extract phenolic compounds from artichoke (Cynara cardunculus L. var. scolymus (L.) Hegi, cultivar "Campagnano") heads. The effect of extraction time, temperature, and solvent-to-sample ratio on the profile and content of phenolic acids and flavonoids was investigated. The total phenolic content was 488.13 ± 0.56 mg GAE 100 g-1 dry matter (dm) and total flavonoid content was 375.03 ± 1.49 mg CATeq 100 g-1 dm when the optimum extraction conditions were set. The HPLC analysis showed that caffeoylquinic acid derivatives (i.e., cynarin and 1,5-O-dicaffeoylquinic acid) were the main compounds in globe artichokes. Caffeic and p-coumaric acids were also identified. In regard to flavonoids, only the flavone luteolin-7-O-glucoside was identified.

Phenolic compounds, antioxidant and antibacterial activities of the methanolic extracts of Euphorbia resinifera and Euphorbia echinus

Euphorbia resinifera and Euphorbia echinus are Moroccan endemic plants that are used in traditional medicine for their therapeutic properties. The aim of this research is to study the phenolic compounds content, the antioxidant and the antibacterial activities of two Euphorbiaceae species: Euphorbia resinifera and Euphorbia echinus, which were collected in Béni-Mellal and Agadir regions, respectively. The current study involved determining the concentration of phenolics, flavonoids, and tannins of our spieces using spectrophotometry. To test the antioxidant activity of the two methanolic extracts, the DPPH° technique was used.Also, the antibacterial activity against three microorganisms was tested in vitro. The quantification of phenolic compounds revealed that Euphorbia resiniferamethanolic extract had the highest concentration of phenolics. The methanolic extract of Euphorbia resinifera and Euphorbia echinus showed a higher antioxidant activity than Ascorbic acid.The antibacterial analysis showed a positive correlation between antioxidant activity and total phenolic content. Our extracts exhibited antibacterial activity against Staphylococcus aureus and Bacillus subtilis. In contrast, it had no inhibitory effect on Escherichia coli.

An alternative approach to validation of liquid chromatography-mass spectrometry methods for the quantification of endogenous compounds

Despite the progress in the quantification of xenobiotics, the development and validation of methods designed for endogenous substances still remain challenging due to the natural presence of the analytes in a biological matrix, leading to the inability to obtain a blank sample. Several generally recognized procedures are described to solve this issue, like using surrogate or analyte-depleted matrices or surrogate analytes. However, the workflows used do not always meet the requirements for developing a reliable analytical method or are cost-intensive.This study aimed to design an alternative approach for preparing validation reference samples using authentic analytical standards while preserving the nature of the biological matrix and solving the problem with the inherent presence of analyzed compounds in a studied matrix. The methodology used is based on the standard-addition type procedure. However, unlike the original method, the addition is modified according to a previously measured basal concentration of monitored substances in the pooled biological sample to obtain a predefined concentration in reference samples according to the European Medicines Agency (EMA) validation guideline. The study shows the advantages of described approach on an example of LC-MS/MS analysis of 15 bile acids in human plasma and compares it with other methods commonly used in this field.The method was successfully validated according to the EMA guideline with lower limit of quantification of 5 nmol/L and linearity in the range of 5 - 2000 nmol/L. Finally, the method was used in a metabolomic study on a cohort of pregnant women (n = 28) to confirm intrahepatic cholestasis, the major liver disease observed in pregnancy.

Chemometric analysis and quantification of major anti-cancer activity compounds from leaves of soursop (Annona muricata Linn.)

Chemometric analysis and quantification of major anti-cancer activity compounds from leaves of soursop (Annona muricata Linn.)

Chemical Composition, Antioxidant Activity, and Antibacterial Activity of Black Poplar Buds’ Hydroalcoholic Macerates from Dobrogea Area

Black poplar buds have high contents of many compounds with therapeutic potential, which are useful in cosmetics and the treatment of various dermatitis, respiratory diseases, etc. The aim of this study was to identify and exploit the local plant resources with biologically active properties from the Dobrogea area, Romania. For this purpose, materials were collected from the mentioned area, and macerates of black poplar were prepared in order to evaluate their qualities as antioxidant and antimicrobial agents. Three different black poplar buds’ hydroalcoholic macerates were analyzed by the Folin–Ciocâlteau method to estimate the total content of phenolic compounds, by the HPLC-DAD method for identification and quantification of the main bioactive compounds and by the DPPH radical scavenging method to evaluate the antioxidant activity. All hydroalcoholic macerates showed high concentrations of phenolic compounds, the main individual compounds being gallic acid, chlorogenic acid, cinnamic acid, and methyl gallic acid. The antioxidant activity of the black poplar buds’ hydroalcoholic macerates, evaluated by the DPPH radical scavenging test, showed high values, between 496 and 1200 mg GAE /100 g d.w. The Cd, Cu, Zn, Ni, and Pb concentrations released in dry poplar buds, determined by AAS, were below the detection limits. Hydroalcoholic macerates of black poplar were tested against two groups of gram-positive bacteria (Enterococcus and Staphylococcus) using an agar well diffusion assay. The in vitro inhibitory activities of the macerates were important and ranged from 8.2–9.4 mm inhibition zones (Staphylococcus) to 8.6 −10 mm inhibition zones (Enterococcus).

Characterization of Phenolic Profile in Milk Obtained by Ewes Fed Grape Pomace: Reflection on Antioxidant and Anti-Inflammatory Status.

The aim of the present work was to evaluate if the use of grape pomace (GP) in the feeding of dairy ewes can improve the content of phenolic compounds (PCs) in the milk and affect the anti-inflammatory and antioxidative status of the milk. For this purpose, 46 ewes were randomly assigned to two groups of 23 animals each: a control group (Ctrl) that received a standard diet and an experimental group (GP+), whose diet was been formulated with 10% GP on a dry matter (DM) basis. At the end of the 60 days of the trial, from 10 ewes selected randomly from each group, individual milk samples were collected and analyzed for the identification and the quantification of phenolic compounds through an ultra-high-performance liquid chromatography system, and milk anti-inflammatory and antioxidative status were evaluated by enzyme-linked immunosorbent assay, determining the activity of GPx and CAT and the levels of IL-1 and TNFα. In addition, gelatinolytic activity of Type IV collagenases (MMP-2/MMP-9) was evaluated by the fluorometric method and zymographic approach. The results obtained showed that the diet with GP affects the phenolic profile of milk, inducing milk enrichment of phenolic compounds without, however, having a significant impact on milk antioxidant and inflammatory status. However, a lower activity of MMP-9 was found in GP+ milk. The use of the molecular docking approach showed the ability of luteolin to approach the catalytic pocket of the enzyme, interfering with the recruitment of the substrate, and therefore, slowing down their hydrolytic activity.

Separation and quantification of Tartrazine (E102) and Brilliant Blue FCF (E133) in green colored foods and beverages

In the present study we investigated the capacities of a panel of 25 solid sorbents represented by layered structures, inorganic oxides and hydroxides, and phyllosilicates, to effectively remove in high yield Tartrazine (E102) and Brilliant Blue FCF (E133) from aqueous solutions, and more notable, green colored food matrices. Quantification of the title compounds have been achieved by HPLC-DAD analyses. Contents of E102 and E133 in real samples were in the range 1.3–36.5 μg/mL and 1.0–20.1 μg/mL, respectively. After a treatment of 1 min., in most cases a complete bleaching of solutions and deep coloring of the solid phase was recorded. The most effective solids to this aim were seen to be aluminium based ayered double hydroxides. In the case of magnesium oxide for E102, and magnesium aluminium D. benzensulfonate SDS 01 H8L and Florisil for E133, a selective adsorption (>99.9 %) of only one dye was observed. The adsorption recorded was strictly dependent on the loading of the sorbent. Related values were 300 mg for the separation of E102 by magnesium oxide from all the five food matrices under investigation, and in the range 200 mg–300 mg for magnesium aluminium D. benzensulfonate SDS 01 H8L and Florisil in the case of E133. The application of Langmuir and Freundlich models suggested that the adsorption may take place in the inner layers of the solids with a favourable thermodynamique outcome. Findings described herein offer the concrete possibility of quantifications of individual dyes in matrices containing more than one food colorant.

HPLC-PDA Method for Quantification of Bioactive Compounds in Crude Extract and Fractions of Aucklandia costus Falc. and Cytotoxicity Studies against Cancer Cells.

Aucklandia costus Falc. (Synonym: Saussurea costus (Falc.) Lipsch.) is a perennial herb of the family Asteraceae. The dried rhizome is an essential herb in the traditional systems of medicine in India, China and Tibet. The important pharmacological activities reported for Aucklandia costus are anticancer, hepatoprotective, antiulcer, antimicrobial, antiparasitic, antioxidant, anti-inflammatory and anti-fatigue activities. The objective of this study was the isolation and quantification of four marker compounds in the crude extract and different fractions of A. costus and the evaluation of the anticancer activity of the crude extract and its different fractions. The four marker compounds isolated from A. costus include dehydrocostus lactone, costunolide, syringin and 5-hydroxymethyl-2-furaldehyde. These four compounds were used as standard compounds for quantification. The chromatographic data showed good resolution and excellent linearity (r2 ˃ 0.993). The validation parameters, such as inter- and intraday precision (RSD < 1.96%) and analyte recovery (97.52-110.20%; RSD < 2.00%),revealed the high sensitivity and reliability of the developed HPLC method. The compounds dehydrocostus lactone and costunolide were concentrated in the hexane fraction (222.08 and 65.07 µg/mg, respectively) and chloroform fraction (99.02 and 30.21 µg/mg, respectively), while the n-butanol fraction is a rich source of syringin (37.91 µg/mg) and 5-hydroxymethyl-2-furaldehyde (7.94 µg/mg). Further, the SRB assay was performed for the evaluation of anticancer activity using lung, colon, breast and prostate cancer cell lines. The hexane and chloroform fractions show excellent IC50 values of 3.37 ± 0.14 and 7.527 ± 0.18 µg/mL, respectively, against the prostate cancer cell line (PC-3).

Straightforward quantification of endogenous steroids with liquid chromatography–tandem mass spectrometry: Comparing calibration approaches

Different calibration strategies are used in liquid chromatography hyphenated to mass spectrometry (LC-MS) bioanalysis. Currently, the surrogate matrix and surrogate analyte represent the most widely used approaches to compensate for the lack of analyte-free matrices in endogenous compounds quantification. In this context, there is a growing interest in rationalizing and simplifying quantitative analysis using a one-point concentration level of stable isotope-labeled (SIL) standards as surrogate calibrants. Accordingly, an internal calibration (IC) can be applied when the instrument response is translated into analyte concentration via the analyte-to-SIL ratio performed directly in the study sample. Since SILs are generally used as internal standards to normalize variability between authentic study sample matrix and surrogate matrix used for the calibration, IC can be calculated even if the calibration protocol was achieved for an external calibration (EC). In this study, a complete dataset of a published and fully validated method to quantify an extended steroid profile in serum was recomputed by adapting the role of SIL internal standards as surrogate calibrants. Using the validation samples, the quantitative performances for IC were comparable with the original method, showing acceptable trueness (79%–115%) and precision (0.8%–11.8%) for the 21 detected steroids. The IC methodology was then applied to human serum samples (n = 51) from healthy women and women diagnosed with mild hyperandrogenism, showing high agreement (R2 > 0.98) with the concentrations obtained using the conventional quantification based on EC. For IC, Passing-Bablok regression showed proportional biases between -15.0% and 11.3% for all quantified steroids, with an average difference of -5.8% compared to EC. These results highlight the reliability and the advantages of implementing IC in clinical laboratories routine to simplify quantification in LC-MS bioanalysis, especially when a large panel of analytes is monitored.

Preliminary Phytochemical Screening, Quantification of Bioactive Compounds and Antioxidant Potential of Achyranthes aspera L. Leaf Powder

Plant extracts have been used since ages for the treatment of various ailments in India and their market is growing globally as well. Achyranthes aspera is one such promising plant with known medicinal properties. Qualitative and quantitative phytochemical screening is necessary to evaluate the therapeutic and antioxidant properties of the medicinal plants. In the present study, we evaluated the bioactive chemical constituents and their in vitro antioxidant potential of A. aspera leaf extract in different solvents. The soxhlets of A. aspera leaf powder in different solvents – methanol, ethanol, aqueous and chloroform on phytochemical screening showed positive results for carbohydrates, phenols, tannins, flavonoids, saponins, cardiac glycosides and steroids. Alkaloids and terpenoids are present in all the extracts except in chloroform extract. Total contents phenols (22.282 gm/100 gm) were found higher as compared to total contents of flavonoids, saponins, alkaloids. Moreover, all antioxidant assays confirmed the antioxidant potential of leaves of A. aspera. Thus, A. aspera is rich in phytochemicals that are needed for its antioxidant potential against various disorders.

Proton Transfer Reactions from N2H+ and ArH+ to Small Molecules

AbstractProton transfer reactions (PTR) are important for chemical ionization, especially in trace gas analysis in air. In an effort to extend their application to the analysis of high‐purity inert gases, we studied PTR of N2H+ and ArH+ with various reaction gases, viz. O2, N2, CH4, CO2, NO2, H2O and CH3OH. Oxygen, nitrogen, methane, carbon dioxide, and water undergo non‐dissociative proton transfer, where the original molecule remains intact. PTR to nitrogen dioxide leads to NO+ with release of neutral OH⋅. While PTR from N2H+ to methanol exclusively yields protonated methanol, the reaction with ArH+ exhibits dissociative proton transfer, due to its higher exothermicity. Products include methenium, protonated formaldehyde, and protonated methanol. The latter undergoes a slow secondary reaction to form protonated dimethyl ether. Within error limits, most reactions proceed at or close to collision rate. Quantum chemical calculations provide energetics and reaction pathways. In such highly exothermic proton transfer reactions, identification and quantification of trace compounds require detailed understanding of the fragmentation pathways and branching ratios.

Application of an integrative system (2D PLE×HPLC-PDA) for bioactive compound extraction and online quantification: Advantages, validation, and considerations

Analyzing compounds such as polyphenols in solid samples frequently uses a solid-liquid extraction step. The solid-liquid extraction and analysis integration in a single equipment is not commercially available since several challenges are inherent to this hybridization. In the context of developing more sustainable analytical procedures, innovative techniques are demanded. Given that, this work proposes a new integrative system (2D PLE × HPLC-PDA) and presents its validation for bioactive compound extraction and online quantification, discussing the main advantages and cares that need to be taken. Two food byproducts – passion fruit bagasse and coffee husks – were chosen as solid model samples. The system was configured to perform pressurized liquid extraction (PLE) with periodical automated extract injection in the HPLC, consequently obtaining the online quantification of target compounds from the solid samples. In parallel with the online injections, extract fractions were collected and submitted to offline analysis in which the extraction yield of piceatannol and chlorogenic acid and caffeine were evaluated, respectively, for passion fruit bagasse and coffee husks. The extraction yields obtained by online and offline injections were compared and were significantly equal (p > 0.05). Thus, the 2D PLE × HPLC-PDA system represents a feasible tool to integrate solid sample preparation and chemical analysis of biocompounds in a single and online step.

Using a Smartphone-Based Colorimetric Device with Molecularly Imprinted Polymer for the Quantification of Tartrazine in Soda Drinks.

The present study reports the development and application of a rapid, low-cost in-situ method for the quantification of tartrazine in carbonated beverages using a smartphone-based colorimetric device with molecularly imprinted polymer (MIP). The MIP was synthesized using the free radical precipitation method with acrylamide (AC) as the functional monomer, N,N'-methylenebisacrylamide (NMBA) as the cross linker, and potassium persulfate (KPS) as radical initiator. The smartphone (RadesPhone)-operated rapid analysis device proposed in this study has dimensions of 10 × 10 × 15 cm and is illuminated internally by light emitting diode (LED) lights with intensity of 170 lux. The analytical methodology involved the use of a smartphone camera to capture images of MIP at various tartrazine concentrations, and the subsequent application of the Image-J software to calculate the red, green, blue (RGB) color values and hue, saturation, value (HSV) values from these images. A multivariate calibration analysis of tartrazine in the range of 0 to 30 mg/L was performed, and the optimum working range was determined to be 0 to 20 mg/L using five principal components and a limit of detection (LOD) of 1.2 mg/L was obtained. Repeatability analysis of tartrazine solutions with concentrations of 4, 8, and 15 mg/L (n = 10) showed a coefficient of variation (% RSD) of less than 6%. The proposed technique was applied to the analysis of five Peruvian soda drinks and the results were compared with the UHPLC reference method. The proposed technique showed a relative error between 6% and 16% and % RSD lower than 6.3%. The results of this study demonstrate that the smartphone-based device is a suitable analytical tool that offers an on-site, cost-effective, and rapid alternative for the quantification of tartrazine in soda drinks. This color analysis device can be used in other molecularly imprinted polymer systems and offers a wide range of possibilities for the detection and quantification of compounds in various industrial and environmental matrices that generate a color change in the MIP matrix.

High-throughput quantification of microbial-derived organic acids in mucin-rich samples via reverse phase high performance liquid chromatography.

Organic acids (short chain fatty acids, amino acids, etc.) are common metabolic byproducts of commensal bacteria of the gut and oral cavity in addition to microbiota associated with chronic infections of the airways, skin, and soft tissues. A ubiquitous characteristic of these body sites in which mucus-rich secretions often accumulate in excess, is the presence of mucins; high molecular weight (HMW), glycosylated proteins that decorate the surfaces of non-keratinized epithelia. Owing to their size, mucins complicate quantification of microbial-derived metabolites as these large glycoproteins preclude use of 1D and 2D gel approaches and can obstruct analytical chromatography columns. Standard approaches for quantification of organic acids in mucin-rich samples typically rely on laborious extractions or outsourcing to laboratories specializing in targeted metabolomics. Here we report a high-throughput sample preparation process that reduces mucin abundance and an accompanying isocratic reverse phase high performance liquid chromatography (HPLC) method that enables quantification of microbial-derived organic acids. This approach allows for accurate quantification of compounds of interest (0.01 mM - 100 mM) with minimal sample preparation, a moderate HPLC method run time, and preservation of both guard and analytical column integrity. This approach paves the way for further analyses of microbial-derived metabolites in complex clinical samples.

UPLC-QTOF-MS/MS-based metabolomic approach and gastroprotective effect of two chemotypes of Egletes viscosa (L.) less. against ethanol-induced gastric ulcer in mice.

Egletes viscosa (L.) (macela) is a native wild herb that can be found in different states of northeastern Brazil. The infusions of its flower buds are traditionally used for the treatment of gastrointestinal disorders. E. viscosa possesses two chemotypes (named A and B), distinguishable by the composition of the essential oil from the flower buds. Although there are previous studies of the gastroprotective effect of the isolated constituents of E. viscosa, its infusions have not been investigated yet. The present study aimed to evaluate and compare the chemical composition and the gastroprotective effect of flower bud infusions of E. viscosa from chemotype A (EVCA) and chemotype B (EVCB). Sixteen infusions were brewed with flower buds according to the traditional preparation mode and were analyzed through a UPLC-QTOF-MS/MS based metabolomic approach for determination of their metabolic fingerprints and quantification of bioactive compounds. Afterward, these data were analyzed by chemometric methods (OPLS-DA) for discrimination of the two chemotypes. Additionally, infusions of EVCA and EVCB (50, 100 and 200mg/kg, p.o.) were evaluated on gastric ulcers induced by absolute ethanol (96%, 0.2mL, p.o.) in mice. To elucidate the gastroprotective mechanisms, the effect of EVCA and EVCB on gastric acid secretion and gastric wall mucus was determined and the role of TRPV1 channels, prostaglandins, nitric oxide and KATP channels were assessed. Moreover, the oxidative stress-related parameters and the histological aspects of the stomach tissue were analyzed. The chemotypes can be discriminated from each other using UPLC-QTOF-MS/MS chemical fingerprints. Both chemotypes presented similar chemical compositions, consisting basically of caffeic acid derivatives, flavonoids and diterpenes. The quantification of bioactive compounds demonstrated that chemotype A possesses more ternatin, tanabalin and centipedic than chemotype B. EVCA and EVCB (50, 100 and 200mg/kg, p.o.) significantly decreased the severity of ethanol-induced gastric lesions, as shown by a reduction in histological alterations and leucocyte infiltration in gastric tissue. The gastroprotective mechanism of both infusions involves an antioxidant effect, maintenance of gastric mucus and reduction gastric secretion. Stimulation of endogenous prostaglandins and nitric oxide release, activation of TRPV1 channels, and KATP channels are also involved in the gastroprotection of the infusions. The gastroprotective effect of EVCA and EVCB was equivalent and mediated through antioxidant and antisecretory actions, including the activation of TRPV1 receptors, stimulation of endogenous prostaglandins and nitric oxide, and opening of KATP channels. The presence of caffeic acid derivatives, flavonoids and diterpenes in both infusions is involved in mediating this protective effect. Our findings support the traditional use of infusions of E. viscosa for gastric disorders regardless of the chemotype.

Phytochemical Screening, Bioactive Compound Quantification by HPLC, and Antifungal Properties of Andrographis paniculata Extracts Against Plant Pathogenic Fungi

The common issues that occurred in plant disease had been studied for many years and fungi were determined as the major microbial agent that typically reduces the crop yield quality and economic losses. The increasing of antimicrobial resistance fungi and chemical fungicides has urged the development of new antifungal agents from medicinal plants such as Andrographis paniculata which is riches in phytochemical compounds. Hence, this research was carried out to investigate the phytochemical compounds and antifungal properties of a botanical extract derived from A. paniculata to inhibit fungal growth. A maceration technique was used where the extracts were soaked in methanol solvent at room temperature (±28°C) for 72 hours. The prepared samples were studied for phytochemical screening, and it shows the presence of alkaloids, flavonoids, saponins, tannins, and terpenoids. Whereas the quantitative estimation of andrographolide compound was conducted by reversed-phase high performance liquid chromatography (HPLC) method. The antifungal test was tested on Fusarium spp., Colletotrichum spp., Neoscytalidium dimidiatum, and Thielaviopsis paradoxa through agar disc diffusion. The result revealed that it exhibits significant fungal growth inhibition. Therefore, this study presented the versatile use of A. paniculata extract in developing its role as a novel broad-spectrum antifungal agent against plant disease.

Identification and Quantification of Polyphenolic Secondary Metabolites in Stem Bark of Ficus religiosa (Moraceae) Using UPLC-HRMS and RP-HPLC-PDA

F. religiosa bark has been extensively used in traditional medicinal systems, such as Ayurveda, for its health benefits. The aim of this study was to investigate the secondary metabolites (phenolics and flavonoids) of the hydroalcoholic stem-bark extract from F. religiosa because this plant has been proven to have a beneficial effect on health disorders. Therefore, a pilot study was conducted for the identification and quantification of polyphenolic compounds in F. religiosa bark using sophisticated chromatographical techniques such as UPLC-HRMS and RP-HPLC-PDA. Additionally, total flavonoids, total phenolics and the scavenging profile of the bark were studied using a UV spectrophotometer. A total of 23 compounds identified with UPLC-HRMS were mainly phenolic acids, polyphenolics, and flavonoids (flavanols and proanthocyanidins). Among the identified compounds, gallic acid, catechin, epicatechin, epigallocatechin gallate, and ellagic acid were simultaneously quantified (0.031–0.380%) using RP-HPLC-PDA. Thereafter, the study complied by evaluating the total flavonoids (109.15 ± 1.2 mg RuE/g and 33.78 ± 0.86 mg CaE/g), total phenolics (4.81 ± 1.01 mg GaE/g), and scavenging profiles (IC50 13.75 ± 0.12 µg/mL) of the F. religiosa bark. This is the first report on the chemical profiling of F. religiosa bark, which is a necessary step to evaluate its nutraceutical properties, paving the way for possible food application.