Characterisation of volatiles and polyphenols for quality assessment of alcoholic beverages prepared from Corsican Myrtus communis berries

In the present work, the essential oil compositions of Myrtus communis berries from ten Corsican localities were studied and no chemical variability was observed. HS–SPME, GC and GC/MS analysis were carried out for the characterisation of volatile fractions of M. communis berries and two derived commercial alcoholic beverages (liqueur and eau-de-vie). Quantitative variations due to the distillation process were observed between the two beverages. The volatile compositions of Corsican myrtle alcoholic products were characterised by high amounts of monoterpene hydrocarbons and oxygenated monoterpenes with α-pinene and 1,8-cineole as major components. The polyphenolic compositions of the berry extract and the liqueur were also established using HPLC–DAD and LC–MS–MS; high concentrations of flavonol glycosides, flavonols and flavanols were reported.     

Secondary metabolites in a soybean fermentation broth of Paecilomyces militaris

A methanolic extract from a soybean broth fermented by Paecilomyces militaris was analysed by HPLC–PDA–HRMS. It mainly contained mannitol, uracil, adenosine, cordycepin, daizein, genistein, and three new isoflavone glycosides. The isoflavonoids were isolated by preparative HPLC and identified with the aid of HPLC–PDA–HRMS, ¹H NMR, ¹³C, H–H COSY, HMBC, and HMQC NMR analysis as: 7,4′-dihydroxyisoflavone-7-O-(4′′-O-methyl)-β-d-glucopyranoside, 7,4′-dihydroxy-6-methoxyisoflavone-7-O-(4′′-O-methyl)-β-d-glucopyranoside, and 5,7,4′-trihydroxyisoflavone-7-O-(4′′-O-methyl)-β-d-glucopyranoside. The three novel compounds are transformation products of soybean isoflavones. The results of metabolites analysis revealed that the fermentation broth reserved the main functional molecules of soybean and P. militaris or Cordyceps militaris. This suggested that soybean fermented with P. militaris may be able to create a combined healthy food. However, isoflavones are also parts of anti-nutrients of soybean products, and cordycepin is a known antibiotic. This means that the fermentation broth may not suit to be used as normal food. At the same time the safety and bioactivity of the novel methylated glycosides need also further approval.     

Quality and antioxidant activity detection of Crataegus leaves using on-line high-performance liquid chromatography with diode array detector coupled to chemiluminescence detection

An on-line high-performance liquid chromatography–diode array detector combined with chemiluminescence detection (HPLC–DAD–CL) method was developed to evaluate the antioxidant activity and quality of Crataegus leaves derived from different species and habitats. The chromatographic fingerprint and active profiles of Crataegus leaves were simultaneously achieved by HPLC–DAD–CL. Main antioxidants in Crataegus leaves were identified by HPLC–DAD–CL and HPLC/ESI/MS. Data analysis based on the total antioxidant activities of 10 batches of samples indicated that there were obvious differences among samples from different species and habitats. Good precision and reproducibility were obtained for DAD and CL detection with relative standard deviations less than 1.0% for retention time and 4.0% for intra and inter-day variations. The results suggested that the on-line method might provide potential information for the identification and quantitation of active markers, which were directly associated with the quality of herbal medicines.     

Comparison of phenolic composition and antioxidant properties of two native Chilean and one domestic strawberry genotypes

The phenolic composition of extracts from the forms chiloensis and patagonica of Fragaria chiloensis were compared with that of the commercial strawberry Fragaria × ananassa cv. Chandler by high performance liquid chromatography with diode-array detector (HPLC–DAD) and high performance liquid chromatography with electrospray ionization mass spectroscopy detector (HPLC–ESI–MS). The phenolic constituents in the three species were mainly proanthocyanidins, hydrolysable tannins, anthocyanins and flavonol glycosides. In both native strawberry species the main flavonol glycoside was quercetin 3-O-glucuronide and the minor anthocyanins identified were cyanidin-malonyl-glucoside and pelargonidin-malonyl-glucoside. The highest anthocyanin content was found in the commercial red strawberry while ellagic acid was the main phenolic in the native white strawberry. From the methanolic fruit extract of the native Chilean strawberry Fragaria chiloensis ssp. chiloensis f. chiloensis four known antioxidants were isolated by selective fractionation using the bleaching of the free radical scavenger 1,1-diphenyl-2-picrylhydrazyl (DPPH) as the guiding assay. The antioxidant properties (measured as the bleaching of the DPPH radical) were determined for methanol extracts of the three fruits, for fractions of F. chiloensis ssp. chiloensis f. chiloensis, and for the isolated compounds cyanidin-3-glucoside, pelargonidin-3-glucoside, quercetin-3-glucuronide and ellagic acid. This study allows a clear chemical differentiation between the commercial strawberry and the Chilean white strawberry.     

Comprehensive analysis of Phyteuma orbiculare L., a wild Alpine food plant

Plants which have been traditionally eaten by the alpine population may provide new opportunities for agricultural development in mountain regions. In this context we have investigated the chemical composition of Phyteuma orbiculare (Campanulaceae), a perennial herb whose leaves have been eaten as salad by rural populations in Valais (Switzerland). Extracts of different polarities were subjected to comprehensive metabolite profiling using a dereplication platform combining HPLC–PDA-MS, and offline NMR analysis. Twenty-three compounds, including various phenolic glycosides, a new dimeric phenylpropanoid glucoside, saponins, and fatty acids were identified online, or after targeted isolation. Selected phenolic constituents were quantitatively assessed by HPLC–PDA analysis. In addition, substances relevant for nutrition, such as β-carotene, fatty acids, ascorbic acid and minerals were quantified in leaves and flowers. The antioxidant capacity was determined with an ORAC assay, and total phenolic compounds were quantified. Finally, the phytochemical profile was compared to that of the related species P. spicatum, P. hemisphaericum and P. ovatum.     

Chemical composition and antioxidant activities of Myrtus communis L. berries extracts

Myrtle (Myrtus communis L.) berries extracts were prepared with solvents at different polarity (water, ethanol, and ethyl acetate) and analysed using different in vitro tests in order to evaluate their antioxidant properties. Antiradical and total antioxidant activities were measured with DPPH and FRAP tests, respectively. Their ability to protect biological molecules was assessed using the cholesterol and LDL oxidation assays. In addition, phenolic compounds and unsaturated fatty acids composition was analysed by HPLC–DAD and HPLC–MS/MS. Ethanol and water extracts showed the highest amount of extracted compounds, but the highest antiradical and antioxidant activities were found in ethanol and ethyl acetate extracts. These extracts were also the ones with the highest content of phenolic compounds. In addition, our results showed a highly significant correlation between the amount of total phenols and antiradical (R² = 0.9993) or antioxidant activities (R² = 0.9985) in these extracts. HPLC–DAD and HPLC–MS analyses showed significant quantitative and qualitative differences among these three extracts. The ethyl acetate extract had the highest protective effect in assays of thermal (140 °C) cholesterol degradation and Cu²⁺-mediated LDL oxidation, inhibiting the reduction of polyunsaturated fatty acids and cholesterol, and the increase of their oxidative products. These results suggest that because of these properties, myrtle berries could be used in dietary supplements preparations or as food additives.     

Determination of the change of flavonoid components as the defence materials of Citrus unshiu Marc. fruit peel against Penicillium digitatum by liquid chromatography coupled with tandem mass spectrometry

A healthy fruit peel of Citrus unshiu Marc. and one infected by Penicillium digitatum were analysed for flavonoids via high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC–MS/MS) in the positive mode with selected ion monitoring (SIM). Among 16 flavonoid components characterised in C. unshiu Marc., four flavanones and nine flavones were identified for the first time. The identified compounds were quantified by HPLC–UV. To investigate the function of the flavonoids as defence materials, the flavonoid content change of the fruit peel inoculated with P. digitatum was monitored by HPLC. The flavonoid concentration in the infected fruit peel decreased initially after the infection and then gradually increased before finally progressively decreasing.     

Quali-quantitative analysis of flavonoids of Cornus mas L. (Cornaceae) fruits

The methanol extract obtained from the ripe fruits of Cornus mas L. (Cornaceae) have been phytochemically studied. On the basis of HPLC–PDA–MS/MSⁿ analysis eight compounds have been identified as quercetin, kaempferol, and aromadendrin glycosilated derivatives. Three major compounds have been also isolated by Sephadex LH-20 column chromatography followed by HPLC and characterised by NMR spectroscopy. Moreover, LC–PDA–MS analysis of the red pigment revealed the presence of three anthocyanins. The quantitative analysis of all compounds was reported.     

Effects induced by the nodulation with Bradyrhizobium japonicum on Glycine max (soybean) metabolism and antioxidant potential

This study was conducted in order to assess the effect of the inoculation of Glycine max (soybean) with Bradyrhizobium japonicum on the metabolite profile and antioxidant potential of its aerial parts. Extracts containing phenolic compounds and organic acids were analysed by HPLC–DAD and HPLC–UV, respectively. Volatiles profile was determined by HS-SPME/GC–IT-MS. The antioxidant activity of acidic and methanolic extracts was assessed against DPPH. Nodulation caused by B. japonicum led to an increase in the contents of phenolic compounds and organic acids. The same was observed for volatile compounds, with compounds like linalyl acetate, menthyl acetate and α-farnesene being detected only in inoculated plants. The phenolic extracts showed stronger antioxidant capacity than the organic acid extracts. In addition, extracts from plants nodulated with B. japonicum exhibited significantly higher antioxidant activity than control samples. These findings suggest that the inoculation with nodulating B. japonicum strains may be employed to manipulate the content of interesting metabolites in G. max aerial parts.     

Novel acylflavones from Sideritis syriaca ssp. syriaca

In the framework of the detailed phytochemical analysis of the aerial parts of Sideritis syriaca, two novel acylflavones were isolated together with three acetylated flavone glycosides and acylated flavone glycosides. The novel acylflavones were identified as isoscutellarein 7-trans-p-coumarate and apigenin 7-,4′-bis(trans-p-coumarate). Their structures were elucidated by means of UV, 1D and 2D NMR, LC–DAD–MS and confirmed by HR-MS spectroscopy.     

Comparative analyses of seeds of wild fruits of Rubus and Sambucus species from Southern Italy: Fatty acid composition of the oil,total phenolic content,antioxidant and anti-inflammatory properties of the methanolic extracts

Fruit seeds are byproducts from fruit processing. Characterisation of the bioactive compounds present in seeds and evaluation of their potential biological properties is therefore of particular importance in view of a possible valorisation of seeds as a source of health beneficial components. In this work, we have analysed the seeds of Sambucus and Rubus species in order to identify their bioactive components and to determine the antioxidant and anti-inflammatory activities of the extracts. We first analysed their oil content, in order to assess the fatty acid profile and tocopherol content. Moreover, the methanolic extracts of the seeds were analysed for their total phenolic contents and antioxidant capacities. Polyphenols were identified by HPLC–ESI–MS/MS analysis. Furthermore, extracts were evaluated for their inhibitory effects on the production of LPS-induced inflammatory mediators (NO, CCL-20) in RAW 264.7 cells.     

HPLC–PDA–ESI–MS/MS profiling and chemopreventive potential of Eucalyptus gomphocephala DC

An HPLC–PDA–ESI/MS/MS method was developed to identify the phytoconstituents of the EtOAc fraction of Eucalyptus gomphocephala DC. The antioxidant effect of the EtOAc fraction together with its sub-fractions was determined in vitro. The cytotoxicity was evaluated on different cell lines. The EtOAc fraction exhibited strong antioxidant activity, reduced the viability of all cell lines and was more active on MCF-7 and HepG-2 cell lines. Subsequently, the cytotoxicity of the sub-fractions and the isolated compounds were tested on MCF-7, HepG-2. The EtOAc fraction possessed potential antitumour promoting properties. It inhibited the stimulated NO (20%), 5-LOX (48.0%) and COX-2 (49.7%) respectively (at concentration of 20 μg/ml). This study suggests that this fraction is a source of different antioxidant and cytotoxic compounds with potential chemopreventive properties that might prevent different stages of the carcinogenesis process.     

Determination of tetracyclines in pig and other meat samples using liquid chromatography coupled with diode array and tandem mass spectrometric detectors

Two high performance liquid chromatographic methods (HPLC–DAD and LC–MS/MS) were developed to analyze tetracycline (TC) residues in pig meat (pork) samples. The method involved a sample preparation using a solid–liquid extraction (SLE) by McIlvaine buffer, followed by a solid-phase extraction (SPE) clean-up using Strata-XL cartridges. The developed sample clean-up resulted in a selective chromatogram in the HPLC–DAD separation and a reduced matrix effect (ME) in LC–MS/MS analysis. Moreover, HPLC columns packed with core–shell particles were tested for separation, which further enhanced the sensitivity and the selectivity of determinations. The validation of the methods for pig samples was carried out according to European Union 2002/657/EC decision. In addition, validation was also performed for bovine, chicken, and turkey meat samples using HPLC–DAD method. The performance characteristics of determinations were evaluated with both spiked and incurred samples, and were systematically compared. LC–MS/MS technique was found to be more accurate for spiked samples; however, HPLC–DAD method resulted in more reliable concentrations for incurred samples.     

Bioassay-guided isolation and identification of antifungal components from propolis against Penicillium italicum

The present study was aimed at identification of antifungal components against Penicillium italicum from Chinese propolis with bioassay-guided fractionation technique. Propolis ethanolic extract (PEE) was separated and purified by liquid–liquid extraction and thin layer chromatography (TLC) and the most active band was subjected to HPLC–MS/MS to identify the antifungal compounds. The results showed PEE and its fractions had strong antifungal activity against P. italicum. Among the fractions of PEE partitioned by petroleum ether, ethyl acetate, n-butanol and water, ethyl acetate fraction (E-Fr) exhibited the most effective activity against P. italicum. Further bioautographic TLC assay showed Band I, with Rf value of 0.70, had an inhibitive zone, which showed the strongest antifungal activity and completely inhibited the growth of P.italicum at 200 mg/L. Bioactive components found in Band I were further identified as pinobanksin, pinocembrine, chrysin and galangin. This study exhibited Chinese propolis and its main flavonoids was potential natural alternatives for the control of citrus blue mould caused by P.italicum.     

Anthocyanin composition and content of the Vaccinium uliginosum berry

Anthocyanins are polyphenols that are widely distributed in plants, and contribute to the brilliant blue, red or purple colour in leaves, flowers or fruits. Their attractive colours and their abundance in the north-east of China make the berries of Vaccinium uliginosum a new and excellent source of natural pigments. The pigments were extracted from the V. uliginosum berry using 0.5% TFA in ethanol solution, purified by an X-5 resin column and characterised by HPLC–DAD analysis. All of the peak assignments were confirmed by low-resolution electrospray mass spectrometry. The results showed that 11 anthocyanins and 2 flavonols were present in the V. uliginosum berry.     

Chemical composition,anthocyanins, non-anthocyanin phenolics and antioxidant activity of wild bilberry (Vaccinium meridionale Swartz) from Colombia

Berries of Vaccinium meridionale Swartz native to Colombia were analysed for chemical composition, total phenolic content, anthocyanin content, and antioxidant activity. In addition, high-performance liquid chromatography with photodiode array detection (HPLC–DAD) and HPLC-electrospray ionisation tandem mass spectrometry (ESI–MS/MS) were used to determine anthocyanin and phenolic composition. Anthocyanin content was 329.0 ± 28.0 mg cyanidin 3-glucoside equivalents/100 g (fresh weight) FW and total phenolic content was 758.6 ± 62.3 mg gallic acid equivalent/100 g FW. Cyanidin 3-galactoside was the major anthocyanin while the most abundant non-anthocyanin phenolic was chlorogenic acid.     

Phytochemical profiles and inhibitory effect on free radical-induced human erythrocyte damage of Dracaena draco leaf: A potential novel antioxidant agent

The present study reports for the first time the metabolite profile and antioxidant activity of aqueous extract obtained from Dracaena draco L. leaf. Volatiles profile was determined by HS-SPME/GC-IT-MS, with 34 compounds being identified, distributed by distinct chemical classes: 2 alcohols, 5 aldehydes, 16 carotenoid derivatives and 8 terpenic compounds. Carotenoid derivative compounds constituted the most abundant class in leaf (representing 45% of total identified compounds). Phenolics profile was determined by HPLC/DAD and 9 constituents were identified: 2 hydroxycinnamic acid derivatives – 5-O-caffeoylquinic and 3,5-O-dicaffeoylquinic acids; 4 hydroxycinnamic acids – caffeic, p-coumaric, ferulic and sinapic acids and 3 flavonol glycosides – quercetin-3-O-rutinoside, kaempferol-3-O-glucoside and kaempferol-3-O-rutinoside. The most abundant phenolic compound is quercetin-3-O-rutinoside (representing 50.2% of total polyphenols). Organic acids composition was also characterised, by HPLC–UV and oxalic, citric, malic and fumaric acids were determined. Oxalic and citric acids were present in higher amounts (representing 47%, each). The antioxidant potential of this material was assessed by the ability to protect against free radical-induced biomembrane damage, using human erythrocyte as in vitro model. Leaf extract strongly protected the erythrocyte membrane from haemolysis (IC₅₀ of 39 ± 11 μg/ml), in a time- and concentration-dependent manner. This is the first report showing that D. draco leaf is a promising antioxidant agent.     

A rapid method for simultaneous determination of triterpenoid saponins in Pulsatilla turczaninovii using microwave-assisted extraction and high performance liquid chromatography–tandem mass spectrometry

In the present study, a microwave-assisted extraction (MAE) and high performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method was developed for the simultaneous determination of seven triterpenoid saponins in Pulsatilla turczaninovii. Operational conditions of MAE were optimized using a central composite design. Multiple-reaction monitoring was employed for quantification while switching the electrospray ion source polarity between positive and negative modes. Full validation of the method was carried out (linearity, precision, accuracy, limit of detection and limit of quantification). The results indicated that the method was rapid, specific and reliable. The developed method was successfully applied to determine the investigated compounds in nine batches of P. turczaninovii. These samples were collected in different seasons from the Liaoning province and varied greatly. The total triterpenoid saponins content was the highest in April, the seedling stage, and decreased from May to August. The contents of total triterpenoid saponins decreased significantly after the blooming period.     

Polyphenolic profiles detected in the ripe berries of Vitis vinifera germplasm

Polyphenolic profiles in the berry samples of 344 European grape (Vitis vinifera) cultivars were evaluated for two consecutive years. These cultivars represent a diverse collection of V. vinifera germplasm maintained at the USDA-Agricultural Research Service Vitis Clonal Repository in Davis of California, USA. A total of 36 polyphenolic compounds, including 16 anthocyanins, 6 flavonols, 6 flavanols, 6 hydroxycinnamic acids and 2 hydroxybenzoic acids, were identified via HPLC–MS and quantified by HPLC–DAD. The mean contents for anthocyanins, flavanols, flavonols, hydroxycinnamic acids and hydroxybenzoic acids were 0.946 (coloured cultivars), 0.147, 0.043, 0.195 and 0.016 mg g⁻¹ FW, respectively. On average, wine grapes had higher concentrations than had table grapes for all of these compounds except hydroxycinnamic acids. Berry colours affected the total contents of anthocyanins, but not others. Positive correlations (0.151–0.535) were found among these groups of compounds. As expected, these groups of compounds were all negatively correlated with berry weight.     

Immunoprecipitation coupled with HPLC–MS/MS to discover the aromatase ligands from Glycyrrhiza uralensis

Affinity chromatography, applied to discover the enzyme inhibitors, needs special column with target protein and its carrier. Selection of stationary phase and mobile phase needs careful considerations due to the characteristics of proteins. In this study, a method immunoprecipitation (IP) coupled with HPLC-DAD–MS was developed to discover the aromatase ligands from Glycyrrhiza uralensis. An SB-C18 column was employed to separate target compounds without special consideration in mobile phase. Twenty-one compounds, including isolated compounds 4, 7, 8, 10, 11, 13, 15, 18–20, 23 and non-isolated compounds A-J, were found to have good affinity to aromatase by LC–MS. Seven of them (7, 15, 18, 19, 23, D, E) were detected to bind with aromatase in MCF-7 cells by IP coupled with HPLC–MS/MS. Bioassays disclosed aromatase inhibitory activities of the isolated compounds mentioned above, verifying the efficiency of IP coupled with HPLC–MS/MS as a method to screen aromatase ligands.