Papers by Silvia Spinozzi
This thesis reports an integrated analytical and physicochemical approach for the study of natura... more This thesis reports an integrated analytical and physicochemical approach for the study of natural substances and new drugs based on mass spectrometry techniques combined with liquid chromatography. In particular, Chapter 1 concerns the study of Berberine a natural substance with pharmacological activity for the treatment of hepatobiliary and intestinal diseases. The first part focused on the relationships between physicochemical properties, pharmacokinetics and metabolism of Berberine and its metabolites. For this purpose a sensitive HPLC-ES-MS/MS method have been developed, validated and used to determine these compounds during their physicochemical properties studies and plasma levels of berberine and its metabolites including berberrubine(M1), demethylenberberine(M3), and jatrorrhizine(M4) in humans. Data show that M1, could have an efficient intestinal absorption by passive diffusion due to a keto-enol tautomerism confirmed by NMR studies and its higher plasma concentration. In...
Journal of Hepatology, 2017
Clinical and Translational Science, 2017
Obeticholic acid (OCA) is a semisynthetic bile acid (BA) analog and potent farnesoid X receptor a... more Obeticholic acid (OCA) is a semisynthetic bile acid (BA) analog and potent farnesoid X receptor agonist approved to treat cholestasis. We evaluated the biodistribution and metabolism of OCA administered to carbon tetrachloride‐induced cirrhotic rats. This was to ascertain if plasma and hepatic concentrations of OCA are potentially more harmful than those of endogenous BAs. After administration of OCA (30 mg/kg), we used liquid chromatography–mass spectrometry to measure OCA, its metabolites, and BAs at different timepoints in various organs and fluids. Plasma and hepatic concentrations of OCA and BAs were higher in cirrhotic rats than in controls. OCA and endogenous BAs had similar metabolic pathways in cirrhotic rats, although OCA hepatic and intestinal clearance were lower than in controls. BAs' qualitative and quantitative compositions were not modified by a single administration of OCA. In all the matrices studied, OCA concentrations were significantly lower than those of en...
Berberine is a quaternary ammonium salt from the protoberberine group of isoquinoline alkaloids f... more Berberine is a quaternary ammonium salt from the protoberberine group of isoquinoline alkaloids found in such plants as gender Berberis. Berberine is recognised to improve glucose and lipid metabolism disorders and preliminary clinical evidences suggest the ability of berberine to reduce endothelial inflammation improving vascular health, even in patients already affected by cardiovascular diseases, suggesting a possible interesting role of berberine and its metabolites in clinical practice. However, its physicochemical properties, pharmacokinetic, and metabolism are not fully elucidated and contradictory data have been reported. This review provides a summary regarding the pharmacological and biological features of berberine, with a focus on berberine as well as their pharmacologically active metabolites and the different mechanisms underlying their activities in order to clarify the correct use of berberine supplementation, alone or in association with other nutraceuticals, for th...
European Journal of Pharmaceutical Sciences
PLoS ONE, Nov 18, 2013
Background: Curcuma extract exerts a myorelaxant effect on the mouse intestine. In view of a poss... more Background: Curcuma extract exerts a myorelaxant effect on the mouse intestine. In view of a possible use of curcuma extract in motor functional disorders of the gastrointestinal tract, a safety profile study has been carried out in the mouse. Methods: Thirty mice were used to study the in vitro effect of curcuma on gallbladder, bladder, aorta and trachea smooth muscular layers and hearth inotropic and chronotropic activity. The myorelaxant effect on the intestine was also thoroughly investigated. Moreover, curcuma extract (200 mg/Kg/day) was orally administered to twenty mice over 28 days and serum liver and lipids parameters were evaluated. Serum, bile and liver bile acids qualitative and quantitative composition was were also studied. Results: In the intestine, curcuma extract appeared as a not competitive inhibitor through cholinergic, histaminergic and serotoninergic receptors and showed spasmolytic effect on K + induced contraction at the level of L type calcium channels. No side effect was observed on bladder, aorta, trachea and heart when we used a dose that is effective on the intestine. An increase in gallbladder tone and contraction was observed. Serum liver and lipids parameters were normal, while a slight increase in serum and liver bile acids concentration and a decrease in bile were observed. Conclusions: Although these data are consistent with the safety of curcuma extract as far as its effect on the smooth muscular layers of different organs and on the heart, the mild cholestatic effect observed in absence of alteration of liver function tests must be further evaluated and the effective dose with minimal side effects considered.
Digestive and Liver Disease
Analytical and Bioanalytical Chemistry
Berberine (BBR) is a natural alkaloid obtained from Berberis species plants, known for its protec... more Berberine (BBR) is a natural alkaloid obtained from Berberis species plants, known for its protective effects against several diseases. Among the primary BBR metabolites, berberrubine (M1) showed the highest plasma concentration but few and conflicting data are available regarding its concentration in biological fluids related to its new potential activity on vascular cells. A combined analytical approach was applied to study biodistribution of M1 in comparison with BBR. The optimization of sample clean-up combined with a fully validated HPLC-ESI-MS/MS tailored for M1 allows sufficient detectability and accuracy to be reached in the different studied organs even when administered at low dose, comparable to that assumed by human. A predictive human vascular endothelial cell-based assay to measure intracellular xanthine oxidase has been developed and applied to study unexplored activities of M1 alongside other common activities. Results showed that oral M1 treatment exhibits higher plasma levels than BBR, reaching maximum concentration 400-fold higher than BBR (204 vs 0.5 ng/mL); moreover, M1 exhibits higher concentrations than BBR also in all the biological compartments analyzed. Noteworthy, the two compounds follow two different excretion routes: M1 through urine, while BBR through feces. In vitro studies demonstrated that M1 inhibited intracellular xanthine oxidase activity, one of the major sources of reactive oxygen species in vasculature, with an IC50 = 9.90 ± 0.01 μg/mL and reduced the expression of the inflammatory marker ICAM-1. These peculiar characteristics allow new perspectives to be opened up for the direct use of M1 instead of BBR in endothelial dysfunction treatment.
Journal of Pharmacology and Experimental Therapeutics, 2014
We report on the relationship between the structure-pharmacokinetics, metabolism, and therapeutic... more We report on the relationship between the structure-pharmacokinetics, metabolism, and therapeutic activity of semi-synthetic bile acid analogues including 6α-ethyl-3α,7αdihydroxy-5β-cholan-24-oic acid (a selective FXR receptor agonist), 6α-ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid (a specific TGR5 receptor agonist), and 6αethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate (a dual FXR/TGR5 agonist). We measured the main physicochemical properties of these molecules, including ionization constants, water solubility, lipophilicity, detergency, and protein binding. Biliary secretion and metabolism, plasma and hepatic concentrations were evaluated by HPLC-ES-MS/MS in bile fistula rat, and compared with natural analogues chenodeoxycholic, cholic and taurochenodexycholic acid intestinal bacteria metabolism was evaluated in terms of 7αdehydroxylase substrate-specificity in anaerobic human stool culture. The semi-synthetic derivatives detergency, measured in terms of their critical micellar concentration was quite similar to the natural analogues. They were slightly more lipophilic than the corresponding natural analogues, evaluated by their 1-octanol water partition coefficient (LogP), due to the ethyl group in 6 position, which makes these molecules very stable toward bacterial 7dehydroxylation. The hepatic metabolism and biliary secretion were different: 6α-ethyl-3α,7α-dihydroxy-5β-cholan-24-oic acid, as chenodeoxycholic acid, were efficiently conjugated with taurine in the liver and only in this form promptly and efficiently secreted in bile. 6α-ethyl-23(S)-methyl-3α,7α,12α-trihydroxy-5β-cholan-24-oic acid was poorly conjugated with taurine, due to the steric hindrance of the methyl at C23 (S) position, metabolized to the C23 (R) isomer and partly conjugated with taurine. Conversely, 6αethyl-3α,7α-dihydroxy-24-nor-5β-cholan-23-sulfate was secreted in bile unmodified and as 3-glucuronide. Therefore, minor structural modifications profoundly influence the metabolism and biodistribution in the target organs where these analogues exert therapeutic effects by interacting with FXR and/or TGR5 receptors.
Research in Veterinary Science, 2014
Aim of this study was to characterize the effects of an ochratoxin A (181 ± 34 ng/g) contaminated... more Aim of this study was to characterize the effects of an ochratoxin A (181 ± 34 ng/g) contaminated diet on growth performances, blood parameters, systemic cytokine levels, cell stress markers and reactivity of immune system of weaned pigs. Growth performance was not affected by OTA consumption even if OTA levels increased in plasma, kidney and liver. OTA diminished the protein content in the serum and increased levels of TNF-alpha and IL-10 in plasma. HO-1 mRNA, indicative for cells stress, was decreased in the kidney but increased in the liver. Additionally, whole blood of the animals of the OTA-group showed a decreased capacity to respond with cytokine expression (mRNA and protein) to ex vivo challenge with LPS. In conclusion our findings indicate that chronic ingestion with OTA-contaminated feed, even at low level, is hazardous for the animal and virtually for human health, pig being an excellent model for human.
Journal of Natural Products, 2014
Berberine (1) is an alkaloid used widely in the treatment of several diseases. However, its physi... more Berberine (1) is an alkaloid used widely in the treatment of several diseases. However, its physicochemical properties, pharmacokinetics, and metabolism remain unclear, and conflicting data have been reported. In this study, the main physicochemical properties of 1 and its metabolites were evaluated, including lipophilicity, solubility, pKa, and albumin binding. A sensitive HPLC-ESIMS/MS method was developed and validated to identify 1 and its main metabolites in human plasma. This method was used to quantify their levels in the plasma of healthy volunteers and hypercholesterolemic patients following a single dose and chronic administration, respectively. In both cases, berberrubine (2) was found to be the main metabolite. Surprisingly, 2 is more lipophilic than 1, which suggests that this compound tautomerizes to a highly conjugated, electroneutral quinoid structure. This was confirmed by NMR studies. These results indicate that the higher plasma concentration of 2 was a consequence of a more efficient intestinal absorption, suggesting that berberrubine is potentially more pharmacologically active than berberine.
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Papers by Silvia Spinozzi