Photochemical and Oxidative Degradation of Chamazulene Contained in Artemisia, Matricaria and Achillea Essential Oils and Setup of Protection Strategies.

Chamazulene (CA) is an intensely blue molecule with a wealth of biological properties. In cosmetics, chamazulene is exploited as a natural coloring and soothing agent. CA is unstable and tends to spontaneously degrade, accelerated by light. We studied the photodegradation of CA upon controlled exposure to UVB-UVA irradiation by multiple techniques, including GC-MS, UHPLC-PDA-ESI-MS/MS and by direct infusion in ESI-MSn, which were matched to in silico mass spectral simulations to identify degradation products. Seven byproducts formed upon UVA exposure for 3 h at 70 mW/cm2 (blue-to-green color change) were identified, including CA dimers and CA benzenoid, which were not found on extended 6 h irradiation (green-to-yellow fading). Photostability tests with reduced irradiance conducted in various solvents in the presence/absence of air indicated highest degradation in acetonitrile in the presence of oxygen, suggesting a photo-oxidative mechanism. Testing in the presence of antioxidants (tocopherol, ascorbyl palmitate, hydroxytyrosol, bakuchiol, γ-terpinene, TEMPO and their combinations) indicated the highest protection by tocopherol and TEMPO. Sunscreens ethylhexyl methoxycinnamate and particularly Tinosorb® S (but not octocrylene) showed good CA photoprotection. Thermal stability tests indicated no degradation of CA in acetonitrile at 50 °C in the dark for 50 days; however, accelerated degradation occurred in the presence of ascorbyl palmitate.

Plant Essential Oils as Healthy Functional Ingredients of Nutraceuticals and Diet Supplements: A Review.

Essential oils (EOs) are mixtures of volatile molecules endowed with health-promoting biological activities that go beyond their role as aromas and natural preservatives and can be exploited to develop functional foods and diet supplements. Their composition is briefly addressed along with regulatory aspects. The potential health benefit of human diet supplementation with EOs is outlined through a review of the recent literature on available clinical trials and preclinical research concerning EOs activity towards: (1) irritable bowel syndrome; (2) inflammatory bowel disease; (3) regulation of microbiota; (4) gastroprotection; (5) hepatoprotection; (6) protection of the urinary tract and diuresis; (7) management of metabolic disorders including hyperglycemia and hyperlipidemia; (8) anti-inflammatory and pain control; (9) immunomodulation and protection from influenza; and (10) neuroprotection and modulation of mood and cognitive performance. The emerging potential in such activities of selected EOs is given focus, particularly green and black cumin, bergamot, orange, myrtle, peppermint, sage, eucalyptus, lavender, thyme, lemon balm, ginger, and garlic.

Real-time oxygen sensing as a powerful tool to investigate tyrosinase kinetics allows revising mechanism and activity of inhibition by glabridin.

A new method for studying tyrosinase kinetics and inhibition by oxygen sensing is described and matched to the conventional spectrophotometric approach. The stoichiometric ratio of O2 uptake to dopachrome formation was 1.5 ±â€¯0.2 for substrate l-tyrosine and 1.0 ±â€¯0.1 for l-DOPA. With both methods, we reinvestigated mushroom tyrosinase inhibition by glabridin from Glycyrrhiza glabra. The two methods agreed showing mixed-type inhibition for monophenolase and diphenolase activities, at variance with previous literature. Average KI (KSI) values for glabridin were 13.6 ±â€¯3.5 (281 ±â€¯89) nM and 57 ±â€¯8 (1312 ±â€¯550) nM, for monophenolase and diphenolase inhibition, respectively, with IC50 of 80 ±â€¯8 nM and 294 ±â€¯25 nM, respectively, at 1 mM substrate. For reference kojic acid KI (KSI) were 10.9 ±â€¯8 (217 ±â€¯55) µM and 9.9 ±â€¯1.4 (21.0 ±â€¯5.2) µM, for monophenolase and diphenolase, respectively, with respective IC50 of 33 ±â€¯8 µM and 17 ±â€¯3 µM. Glabridin's activity is among the highest in nature, being about three orders of magnitude higher than previously reported.

Automatic Measurement of the Carotid Blood Flow for Wearable Sensors: A Pilot Study.

The assessment of the velocity of blood flowing in the carotid, in modern clinical practice, represents an important exam performed both in emergency situations and as part of scheduled screenings. It is typically performed by an expert sonographer who operates a complex and costly clinical echograph. Unfortunately, in developing countries, in rural areas, and even in crowded modern cities, the access to this exam can be limited by the lack of suitable personnel and ultrasound equipment. The recent availability of low-cost, handheld devices has contributed to solving part of the problem, but a wide access to the exam is still hampered by the lack of expert sonographers. In this work, an automated procedure is presented with the hope that, in the near future, it can be integrated into a low-cost, handheld instrument that is also suitable for self-measurement, for example, as can be done today with the finger oximeter. The operator should only place the probe on the neck, transversally with respect to the common tract of the carotid. The system, in real-time, automatically locates the vessel lumen, places the sample volume, and performs an angle-corrected velocity measurement of the common carotid artery peak velocity. In this study, the method was implemented for testing on the ULA-OP 256 scanner. Experiments on flow phantoms and volunteers show a performance in sample volume placement similar to that achieved by expert operators, and an accuracy and repeatability of 3.2% and 4.5%, respectively.

Virtual Real-Time for High PRF Multiline Vector Doppler on ULA-OP 256.

The recent development of high-frame-rate (HFR) imaging/Doppler methods based on the transmission of plane or diverging waves has proposed new challenges to echographic data management and display. Due to the huge amount of data that need to be processed at very high speed, the pulse repetition frequency (PRF) is typically limited to hundreds hertz or few kilohertz. In Doppler applications, a PRF limitation may result unacceptable since it inherently translates to a corresponding limitation in the maximum detectable velocity. In this article, the ULA-OP 256 implementation of a novel ultrasound modality, called virtual real-time (VRT), is described. First, for a given HFR RT modality, the scanner displays the processed results while saving channel data into an internal buffer. Then, ULA-OP 256 switches to VRT mode, according to which the raw data stored in the buffer are immediately reprocessed by the same hardware used in RT. In the two phases, the ULA-OP 256 calculation power can be differently distributed to increase the acquisition frame rate or the quality of processing results. VRT was here used to extend the PRF limit in a multiline vector Doppler (MLVD) application. In RT, the PRF was maximized at the expense of the display quality; in VRT, data were reprocessed at a lower rate in a high-quality display format, which provides more detailed flow information. Experiments are reported in which the MLVD technique is shown capable of working at 16-kHz PRF, so that flow jet velocities higher up to 3 m/s can be detected.

Accurate blood peak velocity estimation using spectral models and vector doppler.

Ultrasound blood peak velocity estimates are routinely used for diagnostics, such as the grading of a stenosis. The peak velocity is typically assessed from the Doppler spectrum by locating the highest frequency detectable from noise. The selected frequency is then converted to velocity by the Doppler equation. This procedure contains several potential sources of error: the frequency selection is noise dependent and sensitive to the spectral broadening, which, in turn, is affected by the Doppler angle uncertainty. The result is, often, an inaccurate estimate. In this work we propose a new method that removes the aforementioned errors. The frequency is selected by exploiting a mathematical model of the Doppler spectrum that has recently been introduced. When a very large sample volume is used, which includes all the vessel section, the model is capable of predicting the exact threshold to be used without the need of broadening compensation. The angle ambiguity is solved by applying the threshold to the Doppler spectra measured from two different directions, according to the vector Doppler technique. The proposed approach has here been validated through Field II simulations, phantom experiments, and tests on volunteers by using defocused waves to insonify a large region from a linear array probe. A mean error lower than 1% and a mean coefficient of variability lower than 5% were measured in a variety of experimental conditions.

Rapid liquid chromatography-tandem mass spectrometry analysis of 4-hydroxynonenal for the assessment of oxidative degradation and safety of vegetable oils.

A novel method for the UHPLC-MS/MS analysis of (E)-4-hydroxynonenal (4-HNE) is described. The method is based on derivatization of 4-HNE with pentafluorophenylhydrazine (1) or 4-trifluoromethylphenylhydrazine (2) in acetonitrile in the presence of trifluoroacetic acid as catalyst at room temperature and allows complete analysis of one sample of vegetable oil in only 21 min, including sample preparation and chromatography. The method involving hydrazine 1, implemented in an ion trap instrument with analysis of the transition m/z 337→154 showed LOD=10.9 nM, average accuracy of 101% and precision ranging 2.5-4.0% RSD intra-day (2.7-4.1% RSD inter-day), with 4-HNE standard solutions. Average recovery from lipid matrices was 96.3% from vaseline oil, 91.3% from sweet almond oil and 105.3% from olive oil. The method was tested on the assessment of safety and oxidative degradation of seven samples of dietary oil (soybean, mixed seeds, corn, peanut, sunflower, olive) and six cosmetic-grade oils (avocado, blackcurrant, apricot kernel, echium, sesame, wheat germ) and effectively detected increased 4-HNE levels in response to chemical (Fenton reaction), photochemical, or thermal stress and aging, aimed at mimicking typical oxidation associated with storage or industrial processing. The method is a convenient, cost-effective and reliable tool to assess quality and safety of vegetable oils.

Acylated anthocyanins from sprouts of Raphanus sativus cv. Sango: isolation, structure elucidation and antioxidant activity.

Little is known on structure-activity relationships of antioxidant anthocyanins. Raphanus sativus cv Sango sprouts are among the richest sources (270 mg/100 g fresh weight). We isolated from sprouts' juice 9 acylated anthocyanins, including 4 new compounds. All comprise a cyanidin core bearing 3-4 glucose units, multiply acylated with malonic and phenolic acids (ferulic and sinapic). All compounds were equally effective in inhibiting the autoxidation of linoleic acid in aqueous micelles, with rate constant for trapping peroxyl radicals kinh=(3.8 ± 0.7) × 10(4)M(-1)s(-1) at 37 °C. In acetonitrile solution kinh varied with acylation: (0.9-2.1) × 10(5)M(-1)s(-1) at 30 °C. Each molecule trapped a number n of peroxyl radicals ranging from 4 to 7. Anthocyanins bearing sinapic acid were more effective than those bearing the ferulic moiety. Under identical settings, deacylated cyanin, ferulic and sinapic acids had kinh of 0.4 × 10(5), 0.3 × 10(5) and 1.6 × 10(5)M(-1)s(-1) respectively, with n ranging 2-3. Results show the major role of acylation on antioxidant performance.

An improved Doppler model for obtaining accurate maximum blood velocities.

Maximum blood velocity estimates are frequently required in diagnostic applications, including carotid stenosis evaluation, arteriovenous fistula inspection, and maternal-fetal examinations. However, the currently used methods for ultrasound measurements are inaccurate and often rely on applying heuristic thresholds to a Doppler power spectrum. A new method that uses a mathematical model to predict the correct threshold that should be used for maximum velocity measurements has recently been introduced. Although it is a valuable and deterministic tool, this method is limited to parabolic flows insonated by uniform pressure fields. In this work, a more generalized technique that overcomes such limitations is presented. The new approach, which uses an extended Doppler spectrum model, has been implemented in an experimental set-up based on a linear array probe that transmits defocused steered waves. The improved model has been validated by Field II simulations and phantom experiments on tubes with diameters between 2mm and 8mm. Using the spectral threshold suggested by the new model significantly higher accuracy estimates of the peak velocity can be achieved than are now clinically attained, including for narrow beams and non-parabolic velocity profiles. In particular, an accuracy of +1.2±2.5 cm/s has been obtained in phantom measurements for velocities ranging from 20 to 80 cm/s. This result represents an improvement that can significantly affect the way maximum blood velocity is investigated today.

Identification and analysis of isothiocyanates and new acylated anthocyanins in the juice of Raphanus sativus cv. Sango sprouts.

The freeze-dried sprouts' juice of Raphanus sativus (L.) cv. Sango was prepared and analysed for the first time. HPLC analysis of total isothiocyanates, after protein displacement, resulted in 77.8 ± 3.0 µmol/g of dry juice while GC-MS analysis of hexane and acetone extracts showed E- and Z-raphasatin (8.9 and 0.11 µmol/g, respectively) and sulforaphene (11.7 µmol/g), summing up to 20.7 ± 1.7 µmol/g of free isothiocyanates. Sprouts' juice contained an unprecedented wealth of anthocyanins and a new fractionation methodology allowed us to isolate 34 mg/g of acylated anthocyanins (28.3 ± 1.9 µmol/g), belonging selectively to the cyanidin family. Analysis was performed by HPLC-PDA-ESI-MS(n) and extended to deacylated anthocyanins and aglycones, obtained, respectively, by alkaline and acid hydrolysis. This study identified 70 anthocyanins, 19 of which have never been described before and 32 of which are reported here in R. sativus for the first time. Sango radish sprouts are exceptional dietary sources of heath-promoting micronutrients.

Exploiting the lipoic acid structure in the search for novel multitarget ligands against Alzheimer's disease.

Lipoic acid (LA) is a natural antioxidant. Its structure was previously combined with that of the acetylcholinesterase inhibitor tacrine to give lipocrine (1), a lead compound multitargeted against Alzheimer's disease (AD). Herein, we further explore LA as a privileged structure for developing multimodal compounds to investigate AD. First, we studied the effect of LA chirality by evaluating the cholinesterase profile of 1's enantiomers. Then, a new series of LA hybrids was designed and synthesized by combining racemic LA with motifs of other known anticholinesterase agents (rivastigmine and memoquin). This afforded 4, which represents a step forward in the search for balanced anticholinesterase and antioxidant capacities.

Non-natural macrocyclic inhibitors of histone deacetylases: design, synthesis, and activity.

Nonpeptidic chiral macrocycles were designed on the basis of an analogue of suberoylanilide hydroxamic acid (2) (SAHA, vorinostat) and evaluated against 11 histone deacetylase (HDAC) isoforms. The identification of critical amino acid residues highly conserved in the cap region of HDACs guided the design of the suberoyl-based macrocycles, which were expected to bear a maximum common substructure required to target the whole HDAC panel. A nanomolar HDAC inhibitory profile was observed for several compounds, which was comparable, if not superior, to that of 2. A promising cytotoxic activity was found for selected macrocycles against lung and colon cancer cell lines. Further elaboration of selected candidates led to compounds with an improved selectivity against HDAC6 over the other isozymes. Pair-fitting analysis was used to compare one of the best candidates with the natural tetrapeptide apicidin, in an effort to define a general pharmacophore that might be useful in the design of surrogates of peptidic macrocycles as potent and isoform-selective inhibitors.

Anomeric discrimination and rapid analysis of underivatized lactose, maltose, and sucrose in vegetable matrices by U-HPLC-ESI-MS/MS using porous graphitic carbon.

Lactose intolerance is a common condition caused by intestinal lactase deficiency, and a lactose-free diet represents the simplest way to avoid gastrointestinal symptoms. The emerging use of dietary supplements requires analytical tools that are capable of assessing these analytes, particularly for those based on dry herbal extracts that contain lactose together with maltose and sucrose, because of cross-contamination and/or deliberate addition as excipient. Electrospray ionization mass spectrometry (ESI-MS) and MS/MS are valuable detection methods for underivatized disaccharides; however, the absence of distinctive ions and collision-induced dissociation (CID) fragmentation patterns does not allow discrimination of stereoisomers without good chromatographic resolution. We developed an ultrahigh performance liquid chromatography-ESI (U-HPLC-ESI) approach, based on porous graphitic carbon (PGC) columns, working at 5 °C to separate and detect the disaccharides in their anomeric forms as formate adducts obtained directly in-column by eluting with formate buffer/acetonitrile gradient mixtures. Using a Paul trap, we monitored the adducts [M + HCOO](-) at m/z 387 in ESI negative mode (MS(1)) as well as the CID fragment ion [M - H](-) at m/z 341 (MS(2)) and used MS(3) fragment ions at m/z 178 and 161 to confirm disaccharides identity in complex vegetable matrices. Complete resolution of lactose α- and ß-anomers, maltose α- and ß-anomers, and sucrose was obtained with R ≥ 2.0 for all peaks and selectivity α = 1.2 between α- and ß-anomers of lactose. The limits of detection were in the range of 3-7 µg/l (ppb) for the target disaccharides. Because of the rapidity and good anomeric discrimination, the described method represents an alternative tool to investigate the mutarotation phenomenon for reducing disaccharides.

Substrate-controlled and organocatalytic asymmetric synthesis of carbocyclic amino acid dipeptide mimetics.

The asymmetric synthesis of a carbocyclic delta-amino acid representing the P(2)/P(3) subunit of a nonpeptidic truncated peptidomimetic molecule is described relying on two independent approaches.

Analysis of in vitro release through reconstructed human epidermis and synthetic membranes of multi-vitamins from cosmetic formulations.

A convenient method for in vitro investigation of the release of lipid- and water-soluble vitamins from cosmetic formulations was developed. The permeation of (d)-alpha-tocopherol (vitamin E), retinyl acetate (pro-vitamin A), ascorbic acid (vitamin C) and pyridoxine (vitamin B6) through SkinEthic reconstructed human epidermis (RHE), and synthetic polyethersulfone and polycarbonate membranes was studied in vitro using a Franz-type diffusion apparatus, coupled either to a spectrophotometer for continuous reading (dynamic setting) or to HPLC-DAD analysis of the receptor medium (static setting). O/W and W/O emulsions were compared with simple aqueous solutions for their kinetic of vitamins release, to evaluate the influence of the cosmetic formulation on the bioavailability of active ingredients. Results indicate that synthetic membranes offer a limited barrier to the diffusion of vitamins, but may provide information on the release ability of the formulation. Penetration was more effective when water was the external phase of the formulation, i.e. W/O emulsions were less effective in the release of vitamins than O/W emulsion or aqueous solutions. RHE (17 days old) offered a significantly higher barrier to penetration of vitamins, as expected for native human epidermis. The relative ranking in coefficient of permeability (Ps (cm/h)) was: ascorbic acid>pyridoxine>>retinyl acetate>alpha-tocopherol approximately 0, the absolute values depending on the formulation. The method herein described showed to be a practical and convenient tool for the quality-control and efficacy evaluation of cosmetic formulations.

Alzheimer's disease: new approaches to drug discovery.

In this work, we review and comment upon the challenges and the 'quo vadis' in Alzheimer's disease drug discovery at the beginning of the new millennium. We emphasize recent approaches that, moving on from a target-centric approach, have produced innovative molecular probes or drug candidates. In particular, the discovery of endosome-targeted BACE1 inhibitors and mitochondria-targeted antioxidants represents a significant advance in Alzheimer's research and therapy. The case study of the development of rasagiline provides an excellent example to support the validity of the multitarget-designed ligand approach to the search for effective medicines for combating Alzheimer's disease.

Potent inhibition of human phosphodiesterase-5 by icariin derivatives.

Plant extracts traditionally used for male impotence (Tribulus terrestris, Ferula hermonis, Epimedium brevicornum, Cinnamomum cassia), and the individual compounds cinnamaldehyde, ferutinin, and icariin, were screened against phosphodiesterase-5A1 (PDE5A1) activity. Human recombinant PDE5A1 was used as the enzyme source. Only E. brevicornum extract (80% inhibition at 50 microg/mL) and its active principle icariin (1) (IC50 5.9 microM) were active. To improve its inhibitory activity, 1 was subjected to various structural modifications. Thus, 3,7-bis(2-hydroxyethyl)icaritin (5), where both sugars in 1 were replaced with hydroxyethyl residues, potently inhibited PDE5A1 with an IC50 very close to that of sildenafil (IC50 75 vs 74 nM). Thus, 5 was 80 times more potent than 1, and its selectivity versus phosphodiesterase-6 (PDE6) and cyclic adenosine monophosphate-phosphodiesterase (cAMP-PDE) was much higher in comparison with sildenafil. The improved pharmacodynamic profile and lack of cytotoxicity on human fibroblasts make compound 5 a promising candidate for further development.

Design, synthesis, and biological evaluation of pirenzepine analogs bearing a 1,2-cyclohexanediamine and perhydroquinoxaline units in exchange for the piperazine ring as antimuscarinics.

Pirenzepine (2) is one of the most selective muscarinic M(1) versus M(2) receptor antagonists known. A series of 2 analogs, in which the piperazyl moiety was replaced by a cis- and trans-cyclohexane-1,2-diamine (3-6) or a trans- and cis-perhydroquinoxaline rings (7 and 8) were prepared, with the aim to investigate the role of the piperazine ring of 2 in the interaction with the muscarinic receptors. The structural change leading to compounds 3-6 abolished in binding assays the muscarinic M(1)/M(2) selectivity of 2, due to an increased M(2) affinity. Rather, compounds 3-6 displayed a reversed selectivity showing more affinity at the muscarinic M(2) receptor than at all the other subtypes tested.

Simple method for the simultaneous isolation and determination of fumonisin B1 and its metabolite aminopentol-1 in swine liver by liquid chromatography--fluorescence detection.

An analytical method based on high-performance liquid chromatography (HPLC) combined with fluorescence detection (FL) has been developed for the simultaneous determination of fumonisin B1 (FB1) and its totally hydrolized metabolite aminopentol-1 (AP1) in pig liver. The sample preparation is based on a single solid phase extraction (SPE). o-Phthalaldehyde (OPA) was used for pre-column derivatization before the programmed reversed-phase analysis on phenylhexyl column. The developed method shows good repeatibility for inter- and intra-day precision as well as adequate linearity of calibration curves (r2 was 0.9855 for FB1 and 0.9831 for AP1). Average recoveries from the matrix were 93.6% for FB1 and 95.3% for AP1. The limit of quantification (LOQ) in swine liver was 75 microg/kg for FB1 and 42 microg/kg for AP1.