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• Mela Rosa Marchigiana pulp callus cultures produce long-lasting bioactive compounds. • Mela Rosa Marchigiana pulp callus extract showed in vitro antioxidant activity. • Mela Rosa Marchigiana pulp callus extract showed ex vivo antioxidant activity. • Mela Rosa Marchigiana pulp callus extract showed in vivo antioxidant activity. [Display omitted] The Mela Rosa Marchigiana (MRM) pulp callus, kept alive since 2017 by monthly sub-cultures, was investigated and morphologically described. In the present paper, the MRM callus ethanolic Extract (MRME) was prepared from one of the 2022 callus sub-cultures and analysed by GC-MS and GC-FID. The investigation was carried out to test whether a long-term culture could affect either the amount or the bioactivity (or both) of the triterpenes produced by the callus. To this aim, the 2022 extract (MRME 2022) was compared to the extract prepared a few years ago (i.e., MRME 2019). Obtained results showed that the composition of both extracts was almost the same, thus revealing that the callus culture efficiency of triterpenes production was nearly unchanged over time. When analysing the biological effect of the extract, all experimental tests, including cell-free, in vitro , ex vivo , and in vivo assays, confirmed that MRME 2022 significantly protected cells, tissues, or whole organisms from oxidative stress-induced damage. Present results could lead to future studies to test MRME as a possible ingredient of a nutraceutical formulation for healthy aging or for the prevention of oxidative stress-associated pathologies. [ABSTRACT FROM AUTHOR]

• ESI-MS/MS fragmentation pattern of the [M+Na]+ ion of PEs was studied. • Fragment ions useful to detect known and new PEs in J. curcas seed oil were found. • Method is useful to detect PEs contamination of derivatives for food use. • Use of MeOH/H 2 O (85:15) in HPLC-ESI-MS/MS analysis enhanced the [M+Na]+ ion signal. Jatropha curcas L. is an inedible plant whose seed oil is an interesting source for biodiesel production. Seed cake, the main byproduct remaining (about 70% w/w) after the oil extraction process, has a high nutritional value but the presence in Jatropha curcas seed of phorbol esters (PEs), a family of toxic compounds with a tigliane skeleton, prevents application of seed cake and other byproducts (e.g. glycerin) in animal feed without an efficient detoxification. Considering the high toxicity of PEs, it is important to have a sensitive analytical method to evaluate the presence of these compounds in Jatropha curcas derivatives. In this paper we present the study of the ESI-MS/MS fragmentation pattern of the [M+Na]+ ion at m / z 733.5 of the six known PEs, namely Jatropha factors (JFs) C 1 -C 6 , which allowed to tentatively identify a series of characteristic and specific fragment ions useful to reveal the presence of JFs in Jatropha curcas seed oil, distinguish them from each other, and identify new PEs (J1-J4). Moreover, the substitution of the usual acetonitrile/water as mobile phase with a mixture of methanol/water (85:15, v/v) allowed to increase the signal of the sodium adduct of about 50-fold during the HPLC-ESI-MS/MS analysis. [ABSTRACT FROM AUTHOR]

Highlights • Purification of sterols and triperpene dialcohols in olive oils. • Use of a polymeric reversed phase and a silica gel direct phase SPE. • Silica gel SPE column with 10–12 µm particle size. • The improved method is simple, time-saving and reduce the amount of solvents used. • Use of 2,2-dimethoxypropane to maintain constant the activity of the silica gel. Abstract The sterols and triterpene dialcohols composition is an important parameter to assess the authenticity of the high value and prone to adulteration extra virgin olive oil. The official methods used to carry out this analysis are time-consuming, labor-intensive and require a high amount of solvents. In this work a simple and time-saving method, based on two solid phase extraction (SPE) steps was developed. After oil saponification, the unsaponifiable matter was purified by polymeric SPE and then the sterols and triterpene dialcohols were isolated by an in-house packed small particle silica gel SPE and analyzed by GC-FID. Results obtained analyzing a sample of extra virgin olive oil, olive oil and refined olive pomace oil with the proposed method showed a good agreement with those obtained with the International Olive Council (IOC) official method. Thus, the use of the proposed method allows a rapid screening for extra virgin olive oils authentication. [ABSTRACT FROM AUTHOR]

Abstract The aim of this work was the study of the best conditions for obtaining a callus culture from the pulp of Acca sellowiana , and to perform a quali-quantitative analysis of the secondary metabolites yielded by the in vitro callus culture. To this end, callus was induced on both Murashige and Skoog and Gamborg B5 media containing various combinations of growth regulators. Three previously undescribed ursane-type triterpenoids, 2α,3β,6α,23-tetrahydroxy-18α,19α-urs-20-en-28-oic acid, 2α,3β,23-trihydroxy-18α,19α-urs-20-en-28-oic acid and 2α,3β,6β,23-tetrahydroxy-18α,19α-urs-20-en-28-oic acid were isolated from the methanolic extract of A. sellowiana culture and characterized by 1D and 2D NMR experiments. Moreover, the quali-quantitative analysis (ESI-MSn and GC–MS) also showed the presence of β-sitosterol, phloridzin, oleanolic, ursolic, 3β-hydroxy-18α,19α-urs-20-en-28-oic, maslinic, corosolic, 2α,3β-dihydroxy-18α,19α-urs-20-en-28-oic, and tormentic acid. Graphical abstract Unlabelled Image Highlights • Optimal conditions to obtain callus cultures from A. sellowiana pulp were studied. • High content of triterpenic acids was found in A. sellowiana fruit pulp culture. • Three undescribed triterpenoids were characterized by 1D and 2D NMR spectra. • The in vitro production of triterpenic acids was higher than in vivo. [ABSTRACT FROM AUTHOR]

A rapid and sensitive HPLC-APCI-MS/MS method for the determination of cholesterol oxidation products (COPs) in milk powder based foods is reported. The method consists in the direct saponification of the sample and purification of oxysterols by reversed phase C18-SPE followed by HPLC-MS/MS analysis. By this procedure, the extraction and enrichment of oxysterols are combined in a unique step, reducing sample manipulation and the possible formation of artifacts. LOD and LOQ were in the concentration ranges of 2–8 ng g −1 and 8–30 ng g −1 , respectively. The precision (CV%) was in the range 10–36% in fresh samples with a total COPs amount from 212 to 645 ng g −1 and 6–14% for an oxidized sample with a higher amount (3651 ng g −1 ). The recovery ranged from 74 ± 8% for 7-ketocholesterol to 101 ± 12% for 7α-hydroxycholesterol at 200 ng g −1 and from 82 ± 2% for 7-ketocholesterol to 117 ± 10% for 5α,6α-epoxycholesterol at 500 ng g −1 spiked levels, respectively. [ABSTRACT FROM AUTHOR]

Grapevine ( Vitis vinifera L.) is currently used for wine production, herbal medicine and food supplements. In this study we investigated the phenolic composition of grape tendril extract (TVV), because these compounds are probably responsible for the observed biological effects of TVV. We performed our investigation on cells implicated in the inflammatory process and in vascular damage, such as monocyte (U937) and senescent endothelial (HUVEC) cells. We analysed changes in microRNA (miRNA) expression during a pro-inflammatory condition in LPS (lipopolysaccharide) stimulated cells. MiR-126, defined as angiomiR for its ability in the maintenance of endothelial cell functions, and miR-146a, defined as inflamma-miR, for its ability to down-modulate (NF-kB nuclear factor-kB) pro-inflammatory pathway, were analysed along with their targets. TVV extract (40 µg/mL) was able to prevent the LPS stimulus and may play an important role in the prevention of endothelial dysfunction when the accumulation of senescent endothelial cells occurs. [ABSTRACT FROM AUTHOR]

[Display omitted] • An innovative method was developed to obtain Cydonia oblonga pulp callus culture. • Pulp callus extract provides a high quantity of secondary compared to fruit pulp. • Pulp callus extract has antioxidant and genoprotective effects. • Pulp callus extract inhibits α-glucosidase lipase and α-amylase. This study has developed an innovative method for the production of secondary metabolites starting from Cydonia oblonga Mill (quince) pulp callus culture. The qualitative and quantitative content of phenolic and triterpenic acids of quince callus extract were elucidated by GC–MS, GC, and HPLC-DAD-ESI-MSn. The callus extract was rich of 5-O-caffeoylquinic acid (5-CQA), 5-p-coumaroylquinic acid (5-p-CoQA) and maslinic and corosolic acid. Quince callus extract's radical scavenging and antioxidant activity were evaluated by 2,2-diphenyl-1-picrylhydrazyl, 2,2,-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid), and Oxygen Radical Absorbance Capacity methods. The genoprotection was evaluated by gel electrophoresis analysis and quantitative Real-Time PCR. In addition to the good antioxidant activity the quince callus extract is a strong inhibitor of α-glucosidase (IC 50 of 0.25 ± 0.02 mg dw/mL) and lipase (IC 50 of 1.99 ± 0.005 mg dw/mL), but mild inhibitor of α-amylase. Therefore, this work would be significant for the future development of a nutraceutical approach to the management of hyperglycemia and dyslipidemia. [ABSTRACT FROM AUTHOR]

Abstract: The aim of this work was to investigate the products of the reactions between isoeugenol and eugenol with the stable 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical and their role in the radical scavenging mechanism. The reaction of isoeugenol and eugenol with the DPPH radical produced, as evidenced by GC–MS and HPLC–MS, a complex mixture of dimeric species in which dehydrodiisoeugenol and its adducts with methanol (reaction solvent) and dehydrodieugenol were the main reaction products, respectively. The antioxidant activity of dehydrodiisoeugenol, determined by the DPPH method, resulted lower than that of isoeugenol considering both the parameters Effective Concentration (EC50) and Antiradical Efficiency (AE). In particular, due to its very slow kinetic behaviour (), the possible contribution of dehydrodiisoeugenol to the DPPH radical scavenging activity of isoeugenol () was practically negligible. On the contrary, dehydrodieugenol had an antioxidant activity higher than that of eugenol and its lower (85min with respect to 126min) made it possible to contribute to the DPPH radical scavenging activity of eugenol. [Copyright &y& Elsevier]

Highlights: [•] The effects of conventional and vacuum roasting of coffee were compared. [•] The effect of vacuum roasting on acrylamide levels of coffee was investigated. [•] Vacuum roasting afforded medium-roasted coffee with lower acrylamide content. [•] Vacuum roasting did not affect the sensory properties of the medium-roasted coffee. [ABSTRACT FROM AUTHOR]

[Display omitted] • 150 MPa high-pressure homogenization produced nano-sized quercetin-loaded liposomes. • Liposomes were oblong in shape (ca. 30 nm) and had a 42% encapsulation efficiency. • Advanced analytical techniques are required to characterize nano-sized liposomes. • Loaded liposomes exhibited higher antitumor action compared to free quercetin. Quercetin-loaded nano-liposomes were prepared by high-pressure homogenization (HPH) at different pressures (up to 150 MPa) and number of passes (up to 3) to define the best processing conditions allowing the lowest particle size and the highest encapsulation efficiency (EE). The process at 150 MPa for 1 pass was the best, producing quercetin-loaded liposomes with the lowest particle size and 42% EE. Advanced techniques (multi-detector asymmetrical-flow field flow fractionation and analytical ultracentrifugation combined with transmission electron microscopy) were further used for the characterization of the liposomes which were oblong in shape (ca. 30 nm). Results highlight the need for several techniques to study nano-sized, polydisperse samples. The potential of quercetin-loaded liposomes against colon cancer cells was demonstrated. Results prove that HPH is an efficient and sustainable method for liposome preparation and highlight the remarkable role of process optimisation as well as the powerfulness of advanced methodologies for the characterisation of nano-structures. [ABSTRACT FROM AUTHOR]

Biostimulants improve yield, quality, and stress acclimation in crops. In this work, we tested the possibility of using phenolics-rich extracts from spelt (Triticum dicoccum L.) husks to attenuate the effects of salt stress (100–200 mM NaCl) in maize. Two methanolic extracts were prepared from the soluble-conjugated (SC), and the insoluble-bound (IB) phenolic acid fractions of the spelt husk, and their effects were investigated on several stress-associated biochemical parameters, such as proline, lipid peroxidation, H 2 O 2 , GSH levels, and ion content. Results show that SC and IB fractions of husk extracts behaved very differently, no doubt due to their greatly divergent chemical composition, as revealed by both GC-MS and HPLC analyses. The efficacy of treatments in mitigating salt stress was also dose- and timing-dependent. IB, even at the lower concentration tested, was able to recover the performance of stressed plants in terms of growth, photosynthetic pigments content, and levels of salt stress markers. Recovery of shoot growth to control levels and reduction of stress-induced proline accumulation occurred regardless of whether plants were pre-treated or post-treated with IB, whereas only pre-treatment with the higher dose of IB was effective in mitigating oxidative stress. Although in some cases SC and even methanol alone exerted some positive effects, they could also be deleterious whereas IB never was. Overall, results indicate that a polyphenol-containing extract obtained from spelt by-products can behave as biostimulant in maize plants and can mitigate their response to salt stress, by acting on different biochemical targets. Image 1 • Two polyphenol-enriched extracts were prepared from spelt by-products. • Chemical composition of the two extracts was assessed by HPLC-DAD and GC-MS analysis. • Maize plants subjected to salt stress were treated with polyphenol-enriched extracts. • The efficacy of spelt husk extracts as biostimulant on maize plants was evaluated. [ABSTRACT FROM AUTHOR]

In order to improve the economic sustainability of the Jatropha-biofuel chain, seed cake detoxification and utilization of 'non toxic' Jatropa curcas accessions are the main activities pursued with the aim of using J. curcas seed cake in animal feed. Given this growing interest, a robust and reliable method for phorbol esters (PEs) determination is necessary. HPLC-UV is a well-established method to detect and quantify the PEs content in Jatropha seeds and related products, but it seems to be unsuitable for more complex matrices like Jatropha leaves and animal tissues, due to the presence of interfering compounds. The objective of this work was to develop and optimize a LC-MS/MS method for the quantitative determination of PEs in seeds and leaves of J. curcas L. plants from Ghana and Mexico and in liver (as an organ with the function of accumulation) from goats fed with PEs in their diet. The HPLC-UV analysis evidenced five chromatographic peaks in the toxic seed kernels corresponding to the factors C1, C2, C3, C6 and C4-C5, respectively, with a PEs concentration of about 5100μg/g (as TPA equivalent). No PEs related peaks were detected in Mexican kernel seeds while in the case of leaves and liver the analysis was hampered by the presence of interfering compounds. The toxic kernel seed extract was used as a standard solution for the PEs quantitation in leaves and liver samples by LC-MS/MS, with the standard addition method. The most intense MRM transitions used to quantify and qualify the PEs were: 675→311, 693→311, and 293→265 m/z. The LC-MS/MS method with a LOD and a LOQ of 0.07 and 0.21μg/g, respectively, resulted in more sensitivity and selectivity than the HPLC-UV method. All three MRM transitions were present in Ghanaian toxic kernel seed, while no peaks were present in the supposed non-toxic Mexican kernel seed. PEs concentration in the leaves of toxic Ghanaian accession resulted in about 1/10 of that in the kernel, while no PEs peaks were found in the J. curcas leaves from Mexico and in liver samples. [ABSTRACT FROM AUTHOR]

Abstract: In this work, a rapid and reliable purification method based on a single mixed solid phase extraction (SPE) column, for the determination of acrylamide in roasted coffee by liquid chromatography–tandem mass spectrometry, was developed. Deuterium labelled d 3-acrylamide was used as internal standard. Acrylamide was extracted by 10mL of water and the extract purified by a single SPE column consisting of 0.5g of an in-house prepared mixture of C18, strong cation (SCX) and anion exchange (SAX) sorbents in the ratio 2/1.5/1.5 (w/w/w). The amount of the three sorbents was optimised in order to eliminate the main interfering compounds present in coffee extracts, such as melanoidins, trigonelline, chlorogenic acids and caffeine. The SPE procedure was very simple and consisted of pushing 1mL of an aqueous coffee extract through the SPE column followed by 1mL of water which was collected for the analysis. The method was tested on six samples of roasted coffee of different composition and roasting level. The repeatability of the method, expressed as relative standard deviation (n =6), was lower than 5%. The recovery of acrylamide at three spiked levels ranged from 92% to 95%. The limits of detection (LOD) and quantitation (LOQ) were 5 and 16μgkg−1, respectively. [Copyright &y& Elsevier]

The reaction between primary amines (RNH2) and diaryl diazomethanes (Ar2CN2) in the presence of catalytic amounts of the complex [RuCl(η5-C5H5)(PPh3)2] (1), in chloroform at 60 °C, generally affords mixtures of imines Ar2C&z.dbnd6;NR as main product and amines Ar2CHNHR. Whereas Ar2CHNHR are formed by the expected carbene insertion into the N&z.sbnd;H bond of the substrate, the generation of Ar2C&z.dbnd6;NR is unprecedented. The carbene intermediate [RuCl(&z.dbnd6;CAr2)(η5-C5H5)(PPh3)] seems to be the key-species involved in the formation of both products. [Copyright &y& Elsevier]