1,721,193 research outputs found
La versatilità del triplo quadrupolo: applicazioni della tecnica precursor ion scanning
No full textAn ocfentanil-related death case: UHPLC-MS analysis of the drug and its metabolites
No full textOcF is a potent synthetic opioid, structurally analogue of fentanyl, abused as a new psychoactive substance in the recreational drug scenario. This work deals with a drug-related fatality involving OcF. In order to obtain a diagnosis of death, a forensic-based method involving anamnestic, clinical and circumstantial, anatomo-pathological and toxicological criteria was used. It was known that the deceased person had a substantial anamnestic history of drug addiction when in life, which may direct the diagnosis towards a death correlate with a chemical cause. As far as the anatomo-pathological findings are concerned, limbs lesions were found to be significant and consisted with a narcotic substance injection. Indeed, the toxicological investigations performed on the brown powder and subsequently on the biological specimens, taken from the body of the deceased person, has shown the presence of OcF in the femoral and cardiac blood, in urine, bile, brain, liver, lung and kidney, together with paracetamol and caffeine. To do this, an ultra-performance liquid chromatography tandem mass spectrometry (UHPLC–MS/MS) based method was developed and validated for OcF quantitation and the simultaneous main metabolites identification. This represents the first reported case in Italy of a fatal outcome related to Ocf and the first time the drug is quantified in tissues. In addition, the toxicological data obtained in the present study, taken together with the well-known potency and danger of the substance and with the congruence between anamnestic, circumstantial and anatomopathological data with the hypothesis of acute narcosis, allow us to support the lethal power of OcF
Vecchie e nuove droghe e loro effetti biologici
No full textAspetti biologici delle sostanze stupefacenti di vecchia e nuova generazione
The “weight” of skin structure and lipidomics on percutaneous absorption of xenobiotics
No full textMolecules can successfully penetrate and permeate skin barrier in relation to their physicochemical characteristics, such as partition coefficient and molecular weight. Nevertheless, skin absorption process is also affected by the highly variable characteristics of skin and by the integrity its upper layers. This latter aspect is particularly critical for ex-vivo skin samples used in in vitro permeation studies.
Aiming to better understand how the membrane variability can affect the results of in vitro permeation studies, the impact of structural characteristics of human epidermis (weight, thickness, electrical resistance properties, storage time) and lipid composition of stratum corneum on the permeation profiles of model molecules were matter of investigation. In line with OECD guideline on in vitro absorption studies, caffeine (log Pow 0.01) and benzoic acid (log Pow 1.83) were used as models, having different lipophilicity. In vitro skin permeation experiments were performed based on Franz diffusion cell and, as a membrane, human epidermis from different donors. The lipid composition of stratum corneum sheets coming from the same donors was obtained by shotgun lipidomics, an analytical approach in which total lipid extracts of tissues are directly infused into a tandem mass spectrometer (triple quadrupole). This approach allows us to detect (and in some cases semi-quantify) about hundred different molecular species, belonging to at least seven lipid classes. The stratum corneum lipid profile obtained was successfully correlated with permeability data
Carnosine and related dipeptides as quenchers of reactive carbonyl species
No full textCarnosine (beta-alanyl-L-histidine, CAR) and related peptides are histidine-containing dipeptides particularly abundant in excitable tissues such as nervous system and skeletal muscle. Although their biochemical role is still unknown, some evidence indicates that these endogenous compounds can act as quenchers of reactive and cytotoxic carbonyl species (RCS). We firstly reported the structural evidence and ex vivo data supporting this hypothesis. As a first step, we investigated the reaction mechanism of CAR as quencher of alpha,beta-unsaturated aldehydes such as 4-hydroxy-trans-2,3-nonenal (HNE) [1] and acrolein (ACR) [2]. It was found that CAR efficiently detoxifies alpha,beta-unsaturated aldehydes through an auto-catalytic mechanism involving both the beta-alanine group and the nucleophilic imidazole ring, forming a stable Michael adduct between the C3 of the aldehyde and the N of the histidine group. The efficacy of CAR and related peptides such as anserine (ANS) as detoxifying agents of HNE was then demonstrated in biological matrices such as spontaneously oxidized rat skeletal muscle, by detecting the corresponding HNE-Michael adducts by liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS)[3]. The ability of CAR to quench ,β-unsaturated aldehydes was then demonstrated ex vivo, by detecting the HNE Michael adduct and correlated metabolites by liquid chromatography-tandem mass spectrometric (LC-MS/MS) approach, based on the precursor ion scanning technique using a triple-stage quadrupole. In particular, the MS approach was applied to urines from Zucker obese rats, a nondiabetic animal model characterized by obesity and hyperlipidemia, where RCS formation plays a key role in the development of renal and cardiac dysfunction. By this approach, the carnosine-HNE Michael adduct was identified together to the two following His metabolites: His-1,4-dihydroxynonane (His-DHN), His-4-hydroxynonanoic acid (His-HNA)[4]. The biological effect of CAR as quencher of RCS was then investigated in Zucker rats supplemented with CAR or with its enantiomer stable to carnosinase (beta-alanyl-D-histidine, D-CAR) (30 mg/kg in drinking water) for 24 weeks. CAR was found to greatly reduce obesity-related diseases in obese Zucker rats, significantly restraining the development of dyslipidaemia, hypertension and renal injury. We believe that the biological effects of CAR in Zucker obese rat are mediated by a direct carbonyl quenching mechanism and this hypothesis is firstly sustained by the in vivo capacity of supplemented CAR to quench RCS, as demonstrated by detecting CAR-HNE Michael adduct in the urine of obese rats, following CAR treatment. Further evidence suggesting a RCS-quenching mechanism is that the diseases inhibited by CAR have a common pathogenetic route, i.e. the oxidative/carbonyl stress, which represents the molecular target of CAR itself. The efficacy of D-CAR was found to be superimposable to that of CAR thus excluding a pro-histamine and receptor-mediated mechanism and confirming a RCS quenching mechanism [5].
In conclusion, among the several biological properties so far reported for CAR and related peptides, their ability to quench and detoxify cytotoxic RCS and in particular alpha,beta-unsaturated aldehydes should also be considered not only to better explain the biochemical role of these dipeptides but also to design novel bioactive drugs.
[1] Aldini G et al. Biochem Biophys Res Commun. 298(5):699-706 (2002).
[2] Carini M et al. J Mass Spectrom. 38(9):996-1006 (2003).
[3] Orioli M et al. J Chromatogr B 827(1):109-18 (2005).
[4] Orioli M et al. Anal Chem. 79(23):9174-84 (2007).
[5] Aldini G et al. J Cell Mol Med. 15(6):1339-1354 (2011)
Bioactive histidine dipeptides: adme studies by lc-ms/ms in humans as a tool for developing new derivatives
No full textCarnosine (β-alanyl-L-histidine, CAR) is the archetype of a class of histidine dipeptides (HP) such as balenine (N-β-alanyl-1-methyl-histidine), homocarnosine (N-4-aminobutyryl-l-histidine, HCAR) and anserine (N-β-alanyl-3-methyl-l-histidine, ANS), widely distributed in the vertebrates, and particularly abundant in the skeletal muscles and excitable tissues. Although CAR has been discovered at the beginning of the last century, its biological role is not yet clarified. However, several pharmacological properties have been up to now reported, such as anti-ischemic, anti-aging and hypoglycemic activities, effects on SNC and in particular on behavior, and preventing/protective efficacy towards diseases related to diabetes and metabolic distress syndrome, such as nephropathy and cardiovascular diseases. Hence, CAR is a promising bioactive agent, and its interest is further sustained by considering the lack of acute and chronic toxicity and that it can be assumed by ingesting a portion of beef. However, only few studies regarding the ADME properties of CAR and His derivatives have been addressed, and most of all conducted in rodents, species lack serum carnosinases, which is supposed to be primarily involved in the metabolic fate of CAR in humans. Hence, aim of the present study was first to monitor the plasma and urine levels of HP after an oral intake of CAR or food containing CAR by an high sensitive and specific HPLC-ESI-MS/MS method set-up and applied in this study. The results show that only a negligible amount of carnosine was detected in plasma after ingesting different types of meat, however, in some cases, plasma anserine concentration was significantly increased as well as urinary carnosine and anserine levels. The fact that no CAR was detected in plasma, clearly due to its fast renal clearance, makes difficult to explain the well-known role of this HP (Aldini 2002; Aldini 2002; Carini 2003; Biofactors 2005; Orioli 2005) as quencher of reactive and cytotoxic carbonyl species, generated by lipooxidation, in vivo. Hence, as second step, to gain a deeper insight into the mechanism of action, we used CAR as a model to start a molecular modelling approach for designing and developing novel drug candidates which, maintaining the trapping activity, and safety of the parent compound, will result more resistant to the enzymatic hydrolysis catalysed by serum carnosinases. The lead compound D-carnosine was evaluated, in parallel to L-CAR, for its carbonyl quenching activity and plasma stability in vitro, as well as for its pharmacokinetics profile in rats
Antioxidant and radical scavenging activity of honey in endothelial cell cultures (EA.hy926)
No full textThe therapeutic properties of honey, once considered a form of folk or preventive medicine, are acquiring importance for the treatment of acute and chronic free radical-mediated diseases (atherosclerosis, diabetes and cancer). The aim of this work was to study the protective activity of a honey of multifloral origin, standardized for total antioxidant power and analytically profiled (HPLC-MS) in antioxidants, in a cultured endothelial cell line (EA.hy926) subjected to oxidative stress. Cumene hydroperoxide (CuOOH) was used as free radical promoter. Native honey (1% w/v pH 7.4, 10(6) cells) showed strong quenching activity against lipophilic cumoxyl and cumoperoxyl radicals, with significant suppression/prevention of cell damage, complete inhibition of cell membrane oxidation, of intracellular ROS production and recovery of intracellular GSH. Experiments with endothelial cells fortified with the isolated fraction from native honey enriched in antioxidants, exposed to peroxyl radicals from 1,1-diphenyl-2-picrylhydrazyl (AAPH, 10 mM) and to hydrogen peroxide (H2O2, 50-100 microM), indicated that phenolic acids and flavonoids were the main causes of the protective effect. These results provide unequivocal evidence that, through the synergistic action of its antioxidants, honey by reducing and removing ROS, may lower the risks and effects of acute and chronic free radical induced pathologies in viv
α,β-Unsaturated aldehydes adducts to actin and albumin as potential biomarkers of carbonylation damage
No full textReactive carbonyl species (RCS) generated by lipid peroxidation, leading to protein carbonylation, are involved in several human diseases. Protein carbonylation constitutes one of the best characterised biomarker of oxidative damage to proteins. Albumin and actin have been identified, through different proteomic approaches, as the main protein targets for RCS in plasma and tissues, respectively. By a combined LC-MS/MS and computational approach, we have demonstrated their high reactivity towards alpha,beta-unsaturated aldehydes, and established the stoichiometry of reaction with HNE and acrolein, as well as the amino acid residues more susceptible to carbonyl attack. A new mass spectrometric approach, based on LC-MS/MS analysis of tag HNE/ACR-modified peptides of carbonylated albumin and actin is proposed, and the advantages over the conventional methods for RCS and RCS-adducted protein analyses discusse
Precursor ion scanning approach for identification of new oxidative-carbonyl stress biomarkers in urine
No full text- …
