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Phytochemical Analysis - Current Research Articles



Current research articles: Phytochemistry

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Phytochemical Analysis - published by Wiley

Phytochemical Analysis devoted to the publication of original articles concerning the development, improvement, validation and/or extension of application of analytical methodology in the plant sciences. The spectrum of coverage is broad, encompassing methods and techniques relevant to the detection (including bio-screening), extraction, separation, purification, identification and quantification of compounds in plant biochemistry, plant cellular and molecular biology, plant biotechnology, the food sciences, agriculture and horticulture




Current articles of the journal:



Investigation of the Role of the Calvin Cycle and C1 Metabolism during HCHO Metabolism in Gaseous HCHO-Treated Petunia under Light and Dark Conditions Using 13C-NMR

Introduction It has been shown that formaldehyde (HCHO) absorbed by plants can be assimilated through the Calvin cycle or C1 metabolism. Our previous study indicated that Petunia hybrida could effectively eliminate HCHO from HCHO-polluted air. Objective To understand the roles of C1 metabolism and the Calvin cycle during HCHO metabolism and detoxification in petunia plants treated with gaseous H13CHO under light and dark conditions. Methods Aseptically grown petunia plants were treated with gaseous H13CHO under dark and light conditions. The metabolites generated from HCHO detoxification in petunia were investigated using 13C-NMR. Results [2-13C]glycine (Gly) was generated via C1 metabolism and [U-13C]glucose (Gluc) was produced through the Calvin cycle simultaneously in petunia treated with low-level gaseous H13CHO under light conditions. Generation of [2-13C]Gly decreased whereas [U-13C]Gluc and [U-13C]fructose (Fruc) production increased greatly under high-level gaseous H13CHO stress in the light. In contrast, [U-13C]Gluc and [U-13C] Fruc production decreased greatly and [2-13C]Gly generation increased significantly under low-level and high-level gaseous H13CHO stress in the dark. Conclusion C1 metabolism and the Calvin cycle contributed differently to HCHO metabolism and detoxification in gaseous H13CHO-treated petunia plants. As the level of gaseous HCHO increased, the role of C1 metabolism decreased and the role of the Calvin cycle increased under light conditions. However, opposite changes were observed in petunia plants under dark conditions. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 19 February 2015 | 6:56 am


Quantitative Determination of Secoiridoids and Phenylpropanoids in Different Extracts of Ligustrum Vulgare L. Leaves by a Validated HPTLC–Photodensitometry Method

Introduction The genus Ligustrum (Oleaceae) is distributed in Europe and Asia (south China and Korea), where it is used to prevent hypertension, sore throats, inflammation and diabetes. The main groups of compounds in extracts of Ligustrum vulgare are biologically active secoiridoids and phenylpropanoids. Objectives The aim of the study was primarily the development and validation of a HPTLC–photodensitometry method for separation and determination of secoiridoids (oleacein, oleuropein) and phenylpropanoids (echinacoside) in different extracts prepared from leaves of L. vulgare. A secondary issue was the quantitative screening of oleacein, oleuropein and echinacoside in extracts from leaves collected at different stages of plant growth (from May to September). Methods A HPTLC–photodensitometry method was developed and validated for quantification of oleuropein, oleacein and echinacoside in plant extracts (aqueous and ethanolic extract, decoction, infusion). Silica gel was used as the stationary phase and dichloromethane:methanol:formic acid:water (80:25:1.5:4, v/v/v/v) as the mobile phase. Results The HPTLC–photodensitometry method developed for quantification of oleacein, oleuropein and echinacoside was specific, accurate and precise. The presence of oleacein was detected in aqueous extracts, whereas oleuropein was present, in particular, in ethanolic extracts, decoctions and infusions. Echinacoside was detected in all the extracts prepared. The content of secoiridoids was variable from May to September, whereas the amount of echinacoside increased in this term. Conclusion The developed and validated HPTLC–photodensitometry method allowed performing fast screening of quantitative profiles of oleacein, oleuropein and echinacoside in preparations of privet leaves. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 19 February 2015 | 6:56 am


Discriminating Lamiophlomis rotata According to Geographical Origin by 1H-NMR Spectroscopy and Multivariate Analysis

Introduction Lamiophlomis rotata (Duyiwei) is a folk herbal medicine that traditionally has been used in China as a hemostatic agent. Raw plant materials used for medicinal products from different geographical regions are often inconsistent in chemical composition. Metabolic fingerprinting provides a new approach for distinguishing the geographical origins of L. rotata. Objective To identify metabolites that contribute to the different geographical regions of L. rotata samples. Methods Lamiophlomis rotata metabolomics were performed by 1H-nuclear magnetic resonance (NMR) spectroscopy and multivariate statistical analyses. The L. rotata metabolic profile was prepared for NMR measurements using methanol-d4 solvent. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were applied to analyse the L. rotata 1H-NMR spectroscopy data. Results Nine iridoid glycosides, one flavonoid and three phenylpropanoid glycosides were detected in L. rotata by 1H-NMR spectroscopy. 1H-NMR measurements and multivariate analysis were used to successfully discriminate samples from three different locations. Conclusion The NMR-based analysis of L. rotata is a more comprehensive approach than traditional chromatographic methods. Simple sample preparation, rapidity and reproducibility of are additional advantages of NMR analysis. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 19 February 2015 | 6:55 am


Root Cause Analysis of Quality Defects Using HPLC–MS Fingerprint Knowledgebase for Batch-to-batch Quality Control of Herbal Drugs

Introduction The batch-to-batch quality consistency of herbal drugs has always been an important issue. Objectives To propose a methodology for batch-to-batch quality control based on HPLC–MS fingerprints and process knowledgebase. Methods The extraction process of Compound E-jiao Oral Liquid was taken as a case study. After establishing the HPLC–MS fingerprint analysis method, the fingerprints of the extract solutions produced under normal and abnormal operation conditions were obtained. Multivariate statistical models were built for fault detection and a discriminant analysis model was built using the probabilistic discriminant partial-least-squares method for fault diagnosis. Results Based on multivariate statistical analysis, process knowledge was acquired and the cause–effect relationship between process deviations and quality defects was revealed. The quality defects were detected successfully by multivariate statistical control charts and the type of process deviations were diagnosed correctly by discriminant analysis. Conclusion This work has demonstrated the benefits of combining HPLC–MS fingerprints, process knowledge and multivariate analysis for the quality control of herbal drugs. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 19 February 2015 | 6:55 am


Densitometric Validation and Optimisation of Polyphenols in Ocimum sanctum Linn by High Performance Thin-layer Chromatography

Introduction Ocimum sanctum Linn (Sanskrit: Tulasi; family: Libiaceae), popularly known as holy basil or Ocimum teinufolium, is found throughout the semitropical and tropical parts of India. In Ayurveda, Tulasi has been well known for its therapeutic potentials. Objective To optimise and develop a standard method to quantify seven polyphenols simultaneously by HPTLC. Methods A three-level factor Box–Behnken statistical design was used for optimisation, where extraction time (min), temperature (°C) and methanol:water ratio (% v/v) are the independent variables with polyphenols as the dependent variable. The separation was archived on a silica-gel 60?F254 HPTLC plate using toluene:ethyl acetate:formic acid:methanol (3:3:0.8:0.2?v/v) as the mobile phase. Densitometric analysis of polyphenols was carried out in the absorbance mode at 366?nm. Results The quantification of polyphenols was carried out based on peak area with a linear calibration curve at concentration ranges of 60–240, 20–200, 100–1600, 40–200, 200–1400, 10–160, 200–1400, 100–5000?ng/band for caffeic acid, ellagic acid, rutin, kaempferol, catechin, quercetin, eupalitin and epicatechin respectively. The method was validated for peak purity, precision, accuracy, limit of detection (LOD) and quantification (LOQ). Method specificity was confirmed using the retention factor value and visible spectra correlation of marker compounds. Conclusions A validated HPTLC method was newly developed for simultaneous quantification of seven polyphenols in an Ayurvedic preparation of O. sanctum. The proposed method is simple, precise, specific, accurate, cost-effective, less time consuming and has the ability to separate the polyphenols from other constituents. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 10 February 2015 | 7:17 am


Recognition of Pyrrolizidine Alkaloid Esters in the Invasive Aquatic Plant Gymnocoronis spilanthoides (Asteraceae)

Introduction The freshwater aquatic plant Gymnocoronis spilanthoides (Senegal tea plant, jazmín del bańado, Falscher Wasserfreund) is an invasive plant in many countries. Behavioural observations of pyrrolizidine alkaloid-pharmacophagous butterflies suggested the presence of pyrrolizidine alkaloids in the plant. Objective To determine whether the attraction of the butterflies to the plant is an accurate indicator of pyrrolizidine alkaloids in G. spilanthoides. Methods The alkaloid fraction of a methanolic extract of G. spilanthoides was analysed using HPLC with electrospray ionisation MS and MS/MS. Two HPLC approaches were used, that is, a C18 reversed-phase column with an acidic mobile phase, and a porous graphitic carbon column with a basic mobile phase. Results Pyrrolizidine alkaloids were confirmed, with the free base forms more prevalent than the N-oxides. The major alkaloids detected were lycopsamine and intermedine. The porous graphitic carbon HPLC column, with basic mobile phase conditions, resulted in better resolution of more pyrrolizidine alkaloids including rinderine, the heliotridine-based epimer of intermedine. Based on the MS/MS and high-resolution MS data, gymnocoronine was tentatively identified as an unusual C9 retronecine ester with 2,3-dihydroxy-2-propenylbutanoic acid. Among several minor-abundance monoester pyrrolizidines recognised, spilanthine was tentatively identified as an ester of isoretronecanol with the unusual 2-acetoxymethylbutanoic acid. Conclusions The butterflies proved to be reliable indicators for the presence of pro-toxic 1,2-dehydropyrrolizidine alkaloids in G. spilanthoides, the first aquatic plant shown to produce these alkaloids. The presence of the anti-herbivory alkaloids may contribute to the plant's invasive capabilities and would certainly be a consideration in any risk assessment of deliberate utilisation of the plant. The prolific growth of the plant and the structural diversity of its pyrrolizidine alkaloids may make it ideal for investigating biosynthetic pathways or for large-scale production of specific alkaloids. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 27 January 2015 | 9:47 am


Systematic Separation and Purification of Iridoid Glycosides and Crocetin Derivatives from Gardenia jasminoides Ellis by High-speed Counter-current Chromatography

Introduction Iridoid glycosides and crocetin derivatives are the main bioactive components of Gardenia. The processes of separation of these compounds reported in much of the literature are tedious, time consuming and require multiple chromatographic steps, which results in lower recovery and higher costs. Objective To develop a high-speed counter-current chromatography (HSCCC) method for the systematic separation and purification of iridoid glycosides and crocetin derivatives on a preparative scale from Gardenia. Methods After fractionation using HPD100 column chromatography, n-butanol:ethanol:water (10:1:10, v/v) was selected to purify gardenoside, 6?-hydroxy geniposide and geniposidic acid from fraction A; ethyl acetate:n-butanol:water (2:1.5:3, v/v) was used to isolate geniposide from fraction B; crocin-1, crocin-2, crocin-3 and crocin-4 were purified by hexane:ethyl acetate:n-butanol:water (1:2:1:5, v/v) from fraction C. The head-to-tail elution mode was used with a flow rate of 8.0?mL/min and a rotary speed of 600?rpm. Results After HSCCC isolation, 151.1?mg of gardenoside, 52.2?mg of 6?-hydroxy geniposide and 24.5?mg of geniposidic acid were obtained from 800?mg of fraction A; 587.2?mg of geniposide was obtained from 800?mg of Fraction B; 246.2?mg of crocin-1, 34.2?mg of crocin-2, 24.4?mg of crocin-3 and 24.7?mg of crocin-4 were obtained from 1000?mg of fraction C. Their purities were found by UPLC analysis to be 91.7%, 93.4%, 92.5%, 98.2%, 94.1%, 96.3%, 94.1% and 98.9% respectively. Conclusion The present results demonstrates that the main iridoid glycosides and crocetin derivatives in Gardenia can be obtained efficiently from extracts using HSCCC. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 27 January 2015 | 9:19 am


Non-targeted Molecular Characterisation of a Rose Flower Ethyl Acetate Extract Using Ultra-HPLC with Atmospheric Pressure Photoionisation and Quadrupole Time-of-flight MS/MS

Introduction A non-targeted approach to characterise the phytochemical composition of the flower organ of an original rose cultivar ‘Jardin de Granville’® was developed. Particular attention was paid to the less documented molecular families of intermediate polarity, compared with the polyphenol family (anthocyanins, flavonoids, tannins) and volatile compounds. Objective To develop a molecular fingerprinting method for the rapid qualitative phytochemical characterisation of the rose flower ethyl acetate extract. Material and methods An ultra-HPLC with atmospheric pressure photoionisation (APPI) and quadrupole time-of-flight (QTOF) MS/MS combined with microwave-assisted extraction was carried out for ethyl acetate extracts as an intermediate polarity extraction solvent in order to obtain the most exhaustive extract containing a large range of molecular families. An optimised methodology based on the coupling of the UHPLC and APPI source with a QTOF analyser was developed to characterise the extracted molecules. Results Sixty-one compounds were identified in the extract, covering eight molecular families of intermediate polarity ranging from polyphenols to triglycerides. The presence of flavonoids with anti-oxidant properties and of triterpenoids with potential anti-inflammatory activity was evidenced and cell-wall constituents such as fatty acids, glycolipids, sphingolipids and acylated sterol glycosides were characterised. Some chlorophyll derivatives were also detected. Conclusion The method developed is appropriate for fast phytochemical evaluation of rose ethyl acetate extract. It produced accurate mass and MS/MS spectra, which permitted identification of a wide range of compounds of intermediate polarity. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 27 January 2015 | 8:43 am


LC–MS/MS Quantitative Determination of Tetrapterys mucronata Alkaloids, a Plant Occasionally used in Ayahuasca Preparation

Introduction Tetrapterys mucronata Cav. (Malpighiaceae) is a plant used in some regions of Brazil in the preparation of ayahuasca. Objective To determine the content of the main tryptamine alkaloids in the stem bark of T. mucronata Cav. and assess their possible toxic and hallucinogenic properties based on the doses found in a water decoction that mimics the ayahuasca preparation. Methods Four alkaloids previously described for their toxic and hallucinogenic properties were quantitated by multiple reaction monitoring HPLC combined with electrospray ionisation and tandem MS (HPLC–ESI/MS/MS) in the water decoction and ethanolic extracts from the bark of T. mucronata. Results Exhaustive extraction of the stem barks with ethanol revealed the following alkaloid levels: bufotenine (1) 3.26?±?0.31?mg/g, 5-methoxy-N-methyltryptamine (2) 0.88?±?0.08?mg/g, 5-methoxy-bufotenine (3) 3.07?±?0.22?mg/g and 2-methyl-6-methoxy-1,2,3,4-tetrahydro-?-carboline (4) 0.14?±?0.004?mg/g. The water decoction presented slightly lower levels, ranging between 2.32?±?0.14, 0.50?±?0.04, 1.53?±?0.09 and 0.10?±?0.01?mg/g for (1), (2), (3) and (4) respectively. Conclusions The HPLC–ESI/MS/MS quantitation revealed significant alkaloid levels, in particular for bufotenine and 5-methoxy-bufotenine. As such compounds are known for their toxic and hallucinogenic properties, these results indicate that the consumption of this plant as an ingredient in ayahuasca preparations may present a risk to consumers. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 26 January 2015 | 6:22 am


Trace Determination of Lead, Chromium and Cadmium in Herbal Medicines Using Ultrasound-Assisted Emulsification Microextraction Combined with Graphite Furnace Atomic Absorption Spectrometry

Introduction The World Health Organization (WHO) recommends that medicinal plants should be checked for the presence of heavy metals. A preconcentration and separation technique for trace amounts of heavy metals from plant matrix is necessary in order to increase the sensitivity and precision of their determination. Objective Lead, chromium and cadmium contaminations in herbal medicines were monitored using ultrasound-assisted emulsification microextraction (USAEME) combined with graphite furnace atomic absorption spectrometry (GF–AAS). Methods In this work, the metal ions in the aqueous solution were complexed with ammonium pyrrolidine dithiocarbamate (APDC) and were extracted into 45??L of toluene that was sonically dispersed in the aqueous phase. The emulsion formed was centrifuged and 20??L of separated toluene was injected into a GF–AAS for analysis. Several factors including the kind of extraction solvent and its volume, sample pH, ionic strength and concentration of APDC were optimised. Results The linear dynamic range (LDR) values were in the range of 0.05 to 20?µg/L and the limit of detection values were in the range of 0.002–0.03?µg/L for target heavy metals. Enrichment factors were obtained in the range of 70–500. The precision of the proposed method was ?8% (n?=?5). The obtained amounts of Pb, Cr and Cd in selected herbal medicines were in the standard range, according to the WHO reports. Conclusion The USAEME with GF–AAS procedure was shown to be an efficient, rapid, inexpensive and eco-friendly method for the determination of lead, chromium and cadmium in herbal medicines. Application of the USAEME method leads to an increased extraction efficiency with satisfactory precision in a short time using an extraction solvent volume at the microlitre level. Copyright © 2015 John Wiley & Sons, Ltd.

Posted on 9 January 2015 | 12:23 pm


Qualitative and Quantitative Analysis of Potentilla fulgens Roots by NMR, Matrix-assisted Laser Desorption/Ionisation with Time-of-Flight MS, Electrospray Ionisation MS/MS and HPLC/UV

Introduction Potentilla fulgens is a commonly used folk medicine by natives of northeast India, Nepal and Bhutan and is rich in polyphenolic and triterpene constituents. Objective To identify chemomarkers in the roots of P. fulgens by an interplay of 13C-NMR, matrix-assisted laser desorption/ionisation with time-of-flight (MALDI/TOF) MS, electrospray ionisation (ESI) MS/MS and HPLC/UV. Material and methods The 13C-NMR spectrum of crude methanolic extract was recorded in deuterated dimethyl sulphoxide. For MALDI/TOF/MS analysis, 2,5-dihydroxybenzoic acid was used as the matrix. For determination of chemical constituents, two independent simple isocratic HPLC/UV methods for monomeric/oligomeric flavanols and triterpene acids were developed and validated. Results The 13C-NMR spectrum of the methanolic extract indicated the presence of B-type oligomeric polyphenolics containing mainly epicatechin/catechin (epicat/cat) and epiafzelechin/afzelechin (epiafz/afz) as the monomeric units. Several isobaric monomeric and oligomeric flavanols and triterpenoids were tentatively identified by MALDI/TOF/MS and ESI/MS/MS. Fourteen compounds (four monomeric and five dimeric flavanols and five triterpene acids) were isolated using repeated column chromatography and semi-preparative HPLC, and were quantitated using HPLC/UV. Conclusion It is evident from these analyses that roots of P. fulgens contain flavans, including oligomeric flavanols, as major constituents followed by triterpene acids. The methods described can be applied to other Potentilla species to identify their constituents. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 16 December 2014 | 11:57 am


Non-targeted Metabolomic Profile of Fagus Sylvatica L. Leaves using Liquid Chromatography with Mass Spectrometry and Gas Chromatography with Mass Spectrometry

Introduction Fagus sylvatica L. is one of the most widely distributed broad-leaved tree species in central and western Europe, important to the forest sector and an accurate biomarker of climate change. Objective To profile the beech leaf metabolome for future studies in order to investigate deeper into the characterisation of its metabolic response. Methods Leaf extracts were analysed using LC–MS by electrospray ionisation in negative mode from m/z 100–1700 and GC–MS by electron ionisation in scan mode from m/z 35–800. Results The LC–MS profile resulted in 56 compounds, of which 43 were identified and/or structurally characterised, including hydroxycinnamic acid derivatives, flavan-3-ols and proanthocyanidins, and flavonols. From a second analysis based on GC–MS, a total of 111 compounds were identified, including carbohydrates, polyalcohols, amino acids, organic acids, fatty acids, phenolic compounds, terpenoids, sterols and other related compounds. Many of the compounds identified were primary metabolites involved in major plant metabolic pathways, however, some secondary metabolites were also detected. Some of them play roles as tolerance-response osmoregulators and osmoprotectors in abiotic stress, or as anti-oxidants that reduce the effect of reactive oxygen species and promote many protective functions in plants. Conclusions This study provides a broad and relevant insight into the metabolic status of F. sylvatica leaves, and serves as a base for future studies on physiological and molecular mechanisms involved in biotic or abiotic stress. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 16 December 2014 | 11:57 am


Isolation of Flavonoids and Triterpenoids from the Fruits of Alphitonia Neocaledonica and Evaluation of their Anti-oxidant, Anti-tyrosinase and Cytotoxic Activities

Introduction Alphitonia neocaledonica (Rhamnaceae) is an endemic tree of New Caledonia. Although three flavonoids have been identified in the leaves, the secondary metabolite profile of the fruits has never been investigated. Objective Phytochemical investigation of A. neocaledonica fruits and evaluation of their anti-oxidant, anti-tyrosinase and cytotoxic activities. Methods A hydromethanolic extract was fractionated by liquid–liquid extraction to obtain ethyl acetate and n-butanolic fractions. The ethyl-acetate-soluble part was purified by silica-gel column chromatography and high-performance liquid chromatography (HPLC). The n-butanol-soluble part was fractionated by centrifugal partition extraction (CPE) and the collected fractions were further purified by centrifugal partition chromatography (CPC) and HPLC. The chemical structures of the purified compounds were identified by nuclear magnetic resonance and mass spectrometry. Results Three triterpenoids and one flavonoid were isolated from the ethyl-acetate-soluble part. Fractions enriched in triterpenoids, flavonoids and catechin derivatives were obtained from the n-butanol-soluble part. Gallocatechin and flavonoids were obtained as pure compounds by further CPC and HPLC purification. The n-butanolic-soluble part showed anti-oxidant and anti-tyrosinase activities due to the presence of tannins and gallocatechin. The triterpenoid alphitolic acid showed a moderate cytotoxic activity against KB cell line (median inhibition concentration?=?8.5??M). Conclusions Nine known compounds including three triterpenes, five flavonoids and (+) gallocatechin, as well as a new 3-O-(6-E-feruloyl)-?-D-glucopyranosyl-(1??2)-[?-D-xylopyranosyl-(1??2)-]?-L-rhamnopyranosyl-quercetin, were isolated from A. neocaledonia fruits. The hydromethanolic extract possesses a potential cytotoxic activity due to the presence of triterpenes, and it can also be valuable as a cosmetic ingredient for its anti-oxidant and anti-tyrosinase activities. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 16 December 2014 | 11:57 am


Ultrasound-assisted Extraction of Gymnemic Acids from Gymnema sylvestre Leaves and its Effect on Insulin-producing RINm-5?F ? Cell Lines

Introduction Gymnema sylvestre is an important anti-diabetic medicinal plant, hence it is necessary to study the effective extraction of its active medicinal components. Objective To develop an efficient ultrasound-assisted extraction method for anti-diabetic gymnemic acids from Gymnema sylvestre leaves and measure their effect on insulin-producing RINm-5?F ? cells. Methods Box–Behnken's design and response surface methodology was applied to the ultrasound-assisted extraction of gymnemic acids from Gymnema sylvestre leaves. Analysis of gymnemic acids was carried out by high-performance thin-layer chromatography by converting total gymnemic acids into gymnemagenin by alkali hydrolysis. Effects of extracts on insulin production were tested on cultured, insulin-producing RINm-5?F ? cell lines. Results The point prediction tool of the design expert software predicted 397.9?mg gymnemic acids per gram of the defatted G. sylvestre leaves using ultrasound-assisted extraction, with ethanol at 60?°C for 30?min. The predicted condition shows 93.34% validity under experimental conditions. The ultrasound-assisted extract caused up to about four times more insulin production from RINm-5?F ? cells than extracts obtained from Soxhlet extraction. Conclusions Response surface methodology was successfully used to improve the extraction of gymnemic acids from G. sylvestre leaves. The ultrasound-assisted extraction process may be a better alternative to prepare such herbal extracts because it saves time and may prevent excess degradation of the target analytes. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 3 December 2014 | 11:23 am


Chemical Differentiation and Quality Evaluation of Commercial Asian and American Ginsengs based on a UHPLC–QTOF/MS/MS Metabolomics Approach

Introduction Asian and American ginsengs are widely used medicinal materials and are being used more and more in health products. The two materials look alike but function differently. Various forms of both types of ginseng are found in the market, causing confusion for consumers in their choice. Objective To evaluate the overall quality of commercial Asian and American ginsengs and investigate the characteristic chemical markers for differentiating between them. Methods This article investigated 17 Asian and 21 American ginseng samples using an ultra-HPLC combined with quadrupole time-of-flight MS/MS technique. The data were processed by principal component analysis and orthogonal partial least squared discriminant analysis. Results In the chromatograms, a total of 40 peaks were detected. Among them, six were positively identified, and all of the remainder were tentatively identified. According to statistical results, ginsenosides Rf, Rb2 and Rc together with their isomers and derivatives were more likely to be present in Asian ginsengs, whereas ginsenoside Rb1, pseudoginsenoside F11 and ginsenoside Rd together with their isomers and derivatives tended to be present in American ginsengs. For Asian ginsengs, ginsenoside Ra3 and 20-?-D-glucopyranosyl-ginsenoside-Rf were more likely to be present in forest samples, whereas contents of floralquinquenoside B, ginsenosides Ro and Rc, and zingibroside R1 were higher in sun-dried ginsengs. For American ginseng, wild samples often had more of the notoginsenosides R1 and Rw2 and less of the ginsenosides Rd, Rd isomer and 20 (S)-Rg3 than cultivated samples. Conclusion The method provided important fingerprint information for authentication and evaluation of Asian and American ginsengs from various commercial products. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 2 December 2014 | 7:02 am


The Evaluation of Reduction of Fe(III) in 3-Hydroxy-4-Nitroso-2,7-Naphthalene Disulphonic Medium as an Alternative Ferric Reducing Activity Power Assay

Introduction The growing interest in determination of anti-oxidant capacity through non-labour, effective and less costly methods encouraged the development of the spectrophotometric procedure presented in this study. Objective To investigate the reduction reaction of Fe(III) in 3-hydroxy-4-nitroso-2,7-naphthalenedisulphonic anion (NRS) medium as an alternative ferric reducing activity power (FRAP) assay for determining the total reduction capacity (RC). Materials and methods The absorbance values at 730?nm were used to determine the RC of aqueous extracts of nine Brazilian plants. The results were compared with the values obtained with the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method and with the total polyphenol content (TPC). The RCs of phenolic derivatives, flavonoids, amino acids and other anti-oxidant compounds were determined. Results Paired t-test with RC values obtained with both assays (proposed FRAP and DPPH) showed no statistically significant difference. In addition, the RC values from the proposed FRAP assay are proportional to those found with TPC values (r =0.916). In addition, the conditional reduction potential of the Fe(III)/Fe(II) couple (0.685?V vs NHE (normal hydrogen electrode)) and the molar absorptivities at 730?nm of the Fe(NRS)33? and Fe(NRS)34? complexes (1.88?×?103 and 1.77?×?104?L/cm?×?mol, respectively) were calculated because these values were not available. Conclusion The proposed assay is adequate for determination of the RC of plant extracts, and the results infer that other samples derived from plants (e.g. beers and wines) and even biological samples (e.g. serum and urine) also could be analysed. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 27 November 2014 | 9:07 am


Development and Validation of Liquid Chromatography Combined with Tandem Mass Spectrometry Methods for the Quantitation of Simalikalactone E in Extracts of Quassia amara L. and in Mouse Blood

Introduction Simalikalactone E (SkE) from Quassia amara, has been proved to be a valuable anti-malarial and anti-cancer compound. As SkE is very scarce, methods of quantitation are needed in order to optimise its isolation process and to determine pharmacokinetic data. Objective To validate methods using liquid chromatography coupled to mass spectrometry for the quantitation of SkE in plant extracts and in biological fluids. Methods High- and ultrahigh-performance liquid chromatography (UHPLC) coupled to ion trap mass spectrometry (MS) with single ion monitoring detection and to triple quadrupole-linear ion trap tandem mass spectrometry with multiple reaction monitoring detection methods were developed. Validation procedure was realised according to the International Conference on Harmonisation guideline. Methanol extracts of dried Quassia amara leaves, and mouse-blood samples obtained after various routes of administration, were analysed for SkE. Results Methods were validated and gave similar results regarding the content of SkE expressed per kilogram of dry leaves in the traditional decoction (160?±?12?mg/kg) and in the methanol extract (93?±?2?mg/kg). The recovery of the analyte from mouse blood ranged from 80.7 to 119.8%. Simalikalactone E was only detected using UHPLC–MS/MS (0.2?±?0.03?mg/L) in mouse blood after intravenous injection: none was detected following intraperitoneal or oral gavage administration of SkE. Conclusion The LC–MS methods were used for the quantitation of SkE in plant extracts and in mouse blood. These methods open the way for further protocol optimisation of SkE extraction and the determination of its pharmacokinetic data. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 27 November 2014 | 9:06 am


In Situ Detection and Identification of Hesperidin Crystals in Satsuma Mandarin (Citrus unshiu) Peel Cells

Introduction Hesperidin, a flavonoid known to have important pharmacological effects, accumulates particularly in the peels of satsuma mandarin (Citrus unshiu). Although histochemical studies have suggested that hesperidin forms crystals in some tissues of the Rutaceae and Umbelliferae, there has been no rigorous in situ detection or identification of hesperidin crystals in C. unshiu. Objective To characterise the chemical component of the crystals found in C. unshiu peels using Raman microscopy. Methods Sections of C. unshiu peels were made. The distribution and morphology of crystals in the sections were analysed microscopically. Raman microscopy was used to detect hesperidin in the sections directly. Results The crystals were more abundant in immature peel and were observed particularly in areas surrounding vascular bundles, around the border between the flavedo and albedo layers and just below the epidermal cells. In the morphological analysis by scanning electron microscopy, needle-shaped crystals aggregated and formed clusters of spherical crystals. Spectra obtained by Raman microscopy of the crystals in the peel sections were consistent with those of the hesperidin standard. Conclusion This study showed the detailed distribution of crystals in C. unshiu peels and their main component was identified using Raman microscopy to be hesperidin for the first time. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 6 November 2014 | 10:18 am


Ultrahigh-performance Liquid Chromatography and Mass Spectrometry (UHPLC–LTQ/Orbitrap/MS/MS) Study of Phenolic Profile of Serbian Poplar Type Propolis

Introduction Propolis is a resinous natural substance collected by honeybees from different plant sources. Due to the presence of various phytochemicals, this bee-product exhibits numerous biological activities, including anti-bacterial, anti-viral, anti-inflammatory, anti-oxidant, immunostimulating and anti-tumour effects. As the chemical composition and biological activity of propolis depend on its botanical and geographical origin, searching for new bioactive substances in various types of propolis from unexplored regions is of great importance. Objective The aim of this study is the evaluation of the phenolic profile of poplar propolis samples in order to characterise Serbian propolis, to identify possible new constituents and to specify the phenolic components relevant for differentiation of poplar propolis samples into two subgroups through simultaneous analysis of poplar bud extracts. Methods Ethanolic extracts of propolis and poplar buds were comprehensively analysed using ultrahigh-performance liquid chromatography coupled with hybrid mass spectrometry, which combines the linear trap quadrupole and Orbitrap MS/MS mass analyser together with chemometric methods. Results Extensive fingerprint analysis of Serbian propolis was achieved for the first time. Seventy-five phenolic compounds were detected. Eight of them were identified in propolis for the first time. Pattern-recognition methods applied to the content of ten quantified phenolics verified the existence of two subgroups of propolis, with galangin, chrysin and pinocembrin as the most influential distinguishing factors. Conclusion The phenolic composition of the analysed propolis samples confirm their affiliation to the European poplar type propolis and the existence of two subgroups according to botanical origin. Copyright © 2014 John Wiley & Sons, Ltd.

Posted on 6 November 2014 | 10:17 am





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